Lead is well known environmental pollutant, which can cause toxic effects in multiple organ systems. However, the influence of lead oxide nanoparticles, frequently emitted to the environment by high temperature technological processes, is still concealed. Therefore, we investigate lead oxide nanoparticle distribution through the body upon their entry into lungs and determine the microscopic and ultramicroscopic changes caused by the nanoparticles in primary and secondary target organs.
Adult female mice (ICR strain) were continuously exposed to lead oxide nanoparticles (PbO-NPs) with an average concentration approximately 106 particles/cm3 for 6 weeks (24 h/day, 7 days/week). At the end of the exposure period, lung, brain, liver, kidney, spleen, and blood were collected for chemical, histological, immunohistochemical and electron microscopic analyses.
Lead content was found to be the highest in the kidney and lungs, followed by the liver and spleen; the smallest content of lead was found in brain. Nanoparticles were located in all analysed tissues and their highest number was found in the lung and liver. Kidney, spleen and brain contained lower number of nanoparticles, being about the same in all three organs. Lungs of animals exposed to lead oxide nanoparticles exhibited hyperaemia, small areas of atelectasis, alveolar emphysema, focal acute catarrhal bronchiolitis and also haemostasis with presence of siderophages in some animals. Nanoparticles were located in phagosomes or formed clusters within cytoplasmic vesicles. In the liver, lead oxide nanoparticle exposure caused hepatic remodeling with enlargement and hydropic degeneration of hepatocytes, centrilobular hypertrophy of hepatocytes with karyomegaly, areas of hepatic necrosis, occasional periportal inflammation, and extensive accumulation of lipid droplets. Nanoparticles were accumulated within mitochondria and peroxisomes forming aggregates enveloped by an electron-dense mitochondrial matrix. Only in some kidney samples, we observed areas of inflammatory infiltrates around renal corpuscles, tubules or vessels in the cortex. Lead oxide nanoparticles were dispersed in the cytoplasm, but not within cell organelles. There were no significant morphological changes in the spleen as a secondary target organ. Thus, pathological changes correlated with the amount of nanoparticles found in cells rather than with the concentration of lead in a given organ.
Sub-chronic exposure to lead oxide nanoparticles has profound negative effects at both cellular and tissue levels. Notably, the fate and arrangement of lead oxide nanoparticles were dependent on the type of organs.
Lead is an environmental pollutant with the largest toxicological database  as it has been a known toxicant for thousands of years. Moreover, lead continues to remain a persistent environmental health threat today. Exposure to lead can result in significant adverse health effects in multiple organ systems. It's toxic effects on the nervous, hematological, renal and reproductive systems have been studied extensively and have been documented in detail .
Lead originates from various industrial and/or household sources, and enters the body through food and fluid intakes, as well as by inhalation [3–5]. Children have been shown to be at greatest risk because of their enhanced gastrointestinal absorption of lead (40–50% vs. 10–15% in adults)  and an incompletely developed blood-brain barrier . Lead exposure increases the risk of diminished intelligence, attention deficit, hyperactivity disorder, school failure and criminal behaviour, thus, there is no known safe level of exposure to lead .
Acute lead poisoning is characterized by non-specific symptoms such as abdominal pain (lead colic), joint pain, constipation, anorexia, muscle aches, headaches, decreased libido, sleep disturbance, irritability, fatigue, anemia, nephropathy, confusion, encephalopathy and seizures . Chronic lead poisoning is the consequence of long-term exposure and can be asymptomatic or may be manifested by a spectrum of cardiovascular and neuropsychiatric diseases, and associated with increased risk of death.
Absorbed lead, which is not excreted out of body, is deposited primarily in mineralizing tissues (bones and teeth), which typically store the majority of the body burden of lead, and in soft tissue organs such as the liver, kidneys, lungs, brain, spleen, muscles and heart . At a steady state, about 90% of lead in the human body was found in the skeleton of adults  and 73% in children, respectively . The half-life of lead in adult human blood has been estimated to range between 28 and 36 days . Lead is excreted via the kidneys with a half-life of about 30 days under normal kidney function. Skeletal lead has a half-life of decades , however, may be gradually released into the bloodstream even after exposure has ceased, particularly during physiological or pathological bone demineralization (such as pregnancy or lactation) .
While the overall negative effect of lead is well known, here we focused our investigation on influence of inhalation of lead oxide nanoparticles (PbO-NPs) on several selected organs. We selected nanoparticles with a diameter of less than 100 nm. Such nanoparticles can represent a serious issue for human health, which was previously shown for similarly sized nanoparticles of different chemistry, because of their easy transport through the cellular barriers [10–12]. Moreover, lead oxide nanoparticles are attention-grabbing substantial for further study as this toxic material is frequently emitted to the environment by high temperature technological processes, such as those used in sintering plants , lead smelters  and electric steel plants , where the lead oxide nanoparticles arise as a result of oxidation process of lead vapors due to the presence of oxygen.
As the respiratory track is one of the most common routes of human exposure to NPs , inhalation of lead oxide nanoparticles was selected for detailed examination of the effect on primary and secondary organs, which have been linked to accumulation, processing or metabolizing the toxins. In this study, we exposed female mice to PbO-NPs by inhalation in a whole body inhalation chamber representing physiological form of exposure similar to real conditions, which is in contrast to intratracheal instillation. In two independent experiments, we used approximately the same average mass concentration of lead oxide nanoparticles (121.7 μg PbO/m3; resp. 149.3 μg PbO/m3). Occupational Safety and Health Administration's (OSHA) set a Permissible Exposure Limit (PEL) for lead in workplace air of 50 μg/m3 (8-h time weighted average) . However, in urban areas and some megacities in India or China [17, 18], concentrations of coarse particulate matter PM10 (PM ≤ 10 μm; containing also metals from anthropogenic sources as Pb, Cd, Zn, Bi, Sb, Cu) can reach pollution peaks at >250 μg/m3 . Thus, mass concentration of nanoparticles applied to our experimental animals was approximately on the same level, which is inhaled by inhabitants of megacities during pollution peaks and also corresponding to the exposure of lead workers in industrial areas.
The aim of this in vivo study is to follow the fate of lead oxide nanoparticles upon entry via the lungs, with the main focus on uncovering their localization and microscopic/ultramicroscopic changes caused by these nanoparticles to target soft tissues.
Adult female mice (ICR strain) were obtained from the Animal Facility of the Masaryk University (Brno, Czech Republic) and were allowed to acclimatize to laboratory conditions for at least one week before the inhalation experiments. Commercial feed and water were provided ad libitum.
PbO nanoparticles (PbO-NPs) were generated continuously in situ in a hot-wall tube flow reactor using an evaporation–oxidation–condensation technique in which a ceramic crucible containing a small amount of lead wire was placed inside the ceramic work tube of a vertically orientated furnace (Carbolite TZF 15/50/610). The melted lead was evaporated at the centre of the furnace at a temperature of 830 °C. The metal vapour formed was carried out of the furnace by an inert nitrogen gas stream and diluted with a stream of air, during which the lead was oxidised to lead oxide. Both flow rates were set at 3 L/min by means of mass flow controllers (MFC). Resulting lead oxide nanoparticles were diluted in the second step by a stream of air (20 L/min) and used for whole-body inhalation experiments in exposure chambers.
The distribution of nanoparticles (NPs) with respect to particle concentration was measured directly and continuously using a Scanning Mobility Particle Sizer (SMPS, model 3936 L72, TSI Inc., Shoreview, USA), with a size range of 8–230 nm (Additional file 1: Figure S1).
The long-term stability of PbO-NPs generation was high. The relative standard deviations in median particle diameter (25.8 nm) and total particle concentration (the first experiment: 1.23 × 106 particles/cm3; the second experiment: 0.956 × 106 particles/cm3) were 3.9% and 3.1%, respectively. These values indicate that both the generation of NPs and measurement by SMPS were reproducible.
Mass concentration of generated PbO nanoparticles was calculated by dividing the mass of PbO nanoparticles collected on the filter by the volume of air sample that was passed through the filter. Generated PbO nanoparticles were sampled on nitrocellulose filters (pore size 0.45 μm, diameter 25 mm, Millipore, Bedford, USA). Filters were dissolved in HNO3 using a UniClever microwave mineraliser (Plazmatronika, Wroclaw, Poland) and Pb content of the sample analysed by means of AAS (AAnalyst 600, PerkinElmer Inc., Shelton, CT, USA).
Surface area of the generated PbO-NPs (the first experiment: 4.30 × 103 μm2/cm3; the second experiment: 4.21 × 103 μm2/cm3) was calculated using the SMPS software .
Size and shape of generated PbO nanoparticles were determined by electron microscopy. Immediately after generation at the furnace output, PbO-NPs were collected by electrostatic precipitation using a Nanometer aerosol sampler (model 3089, TSI) on TEM grids (copper S160–4, 3 mm in diameter, 400 mesh grids, Agar Scientific, Electron Technology, Stansted, Essex, UK). The samples were analysed by means of a Magellan 400 L XHR microscope (FEI Company, Hillsboro, OR, USA), operating in the STEM mode. The PbO-NPs observed in the gas phase by SMPS were formed from agglomeration (size range approximately 40–50 nm) of primary 1–3 nm diameter particles (Additional file 1: Figure S2).
The inhalation chamber has been described in our previous study . Adult female mice, with an average weight of approximately 24 g, were continuously exposed to lead oxide nanoparticles for 6 weeks (24 h/day, 7 days/week). Control animals were exposed to the same air as treated animals group without the addition of nanoparticles. Ten biological replicates (individuals) were used for each treatment. Two independent experiments were accomplished.
Animals in the experimental group (Pb) were exposed to an average concentration of 1.23 × 106 particles/cm3; resp. 0.956 × 106 particles/cm3 (mode 25.9 nm, geometric mean diameter 25.9 nm, median 25.8 nm, geometric standard deviation 1.66, average mass concentration 121.7 μg PbO/m3; resp. 149.3 μg PbO/m3). The estimated deposited dose of PbO over the 6-week inhalation period was 0.75 μg (resp. 0.92 μg in the second experiment) of PbO per gram of mouse body weight [22–24]. Detail calculation of deposited dose of PbO nanoparticles can be found in supplementary online material.
At the end of the exposure period, mice were sacrificed by cervical dislocation. The following organs were collected for chemical, histological and electron microscopic analyses: lung, brain, liver, kidney, and spleen.
For histological analysis, organs were fixed in 10% buffered neutral formaldehyde. Following overnight fixation in the fridge, samples were dehydrated using a series of increasing concentrations of ethanol, immersed in xylene and embedded in paraffin wax. Serial histological sections of 5 μm thickness were prepared and stained using standard Hematoxylin-Eosin staining. Selected sections were stained by Periodic Acid–Schiff (PAS) for detection of glycogen, by Green Trichrome for demonstration of connective tissue elements, especially collagen fibers, and by Luxol Fast Blue for visualization of myelin/myelinated axons in nerve tissue. Sections were examined by light microscopy (Leica DM5000 B, Leica Mikrosysteme GmbH, Vienna, Austria) in a blinded fashion. Photomicrographs were taken using a digital colour camera Leica DFC480 (Leica Mikrosysteme GmbH, Vienna, Austria).
After deparaffinization and rehydration, sections were pre-treated in Target Retrieval Solution (pH = 9, DAKO, cat. No. S2367) for 20 min in a 97 °C water-bath. Non-specific secondary antibody binding was inhibited by incubation in a blocking serum for 20 min at room temperature (RT). Slides were incubated with the primary monoclonal antibody (PCNA, cat. No. 931143, full concentration, Invitrogen, USA; neurofilament, NF-M, cat. No. 2H3 DSHB, USA) for 1 h at RT. Secondary antibody was applied for 30 min at RT. The peroxidase-conjugated streptavidin-biotin system (Vectastain Elite ABC-Peroxidase kit, Vector Laboratories, USA) and chromogen substrate diaminobenzidine (DAB, cat. No. K3466, Dako, USA) were used for visualization. Sections were counterstained using Haematoxylin. A negative control was obtained by omitting the primary antibody from the labelling protocol.
For the detection of apoptotic cells, the nuclear DNA fragmentation was labelled in situ by the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL) method, according to the manufacturer's protocol (ApopTag Peroxidase in situ Apoptosis Detection Kit; Chemicon, Temecula, CA, USA). Counterstaining with Haematoxylin was performed. A negative control was obtained by omitting the enzyme from the labelling protocol.
Following deparaffinization and rehydration, sections were pre-treated in Citrate Buffer (pH = 6) for 20 min in a 97 °C water-bath. Primary antibody (Na-K ATPase, cat. No. ab76020, Abcam, UK) was applied for 1 h at room temperature or overnight at 4 °C. Secondary antibody (goat anti-mouse Alexa Fluor® 488, cat. No. A-11001, Thermo Fisher, USA) was applied for 30 min at room temperature. Sections were washed in PBS and coverslipped in Prolong Gold anti-fade reagent containing DAPI (cat. No. P36935, Invitrogen, USA). The photos taken under a fluorescence microscope Leica DM LB2 (Leica Microsystems, Germany) were merged together in Adobe Photoshop 7.0 (Adobe, USA).
Samples of lungs, brain, liver, kidney and spleen were fixed in 3% glutaraldehyde for 24 h, washed three times in 0.1 M cacodylate buffer and post-fixed in 1% OsO4 solution for 1 h. Samples were then dehydrated in ethanol, followed by acetone and embedded in epoxy resin Durcupan. Semithin sections were stained with Toluidine Blue and examined by light microscopy in a blinded fashion. Ultra-thin sections (60 nm thick) were cut using an ultramicrotome Leica EM UC6 (Leica Mikrosysteme GmbH, Vienna, Austria) and placed on formvar-coated nickel grids.
Sections without further contrasting were observed using Morgagni™ 268 TEM (FEI Company, Eindhoven, Netherlands). Photographs were taken using a Veleta CCD camera (Olympus, Münster, Germany).
Elemental composition was determined by X-Energy Dispersive Spectroscopy (X-EDS). Ultra-thin sections were analysed on a TECNAI 120 (FEI Company, Eindhoven, Netherlands), equipped with a calibrated X-EDS Esprit2 (Bruker AXS, Madison, WI, USA). Working parameters were as follows: accelerating voltage of 120 keV with a LaB6 filament saturated with an emission of 3 (scale of 6); condenser aperture of 100 μm apparent diameter without objective diaphragm. As the TEM mode the Nanoprobe one was used allowing a beam size of 70–100 nm. The sample was tilted at 15° solid angle to achieve a take-off angle of 45°. Counting times were a minimum of 120 s, up to 200 s, with a dead-time between 5% and 20%. These parameters optimised the analytical time regarding the degradation of the sample by the electron beam.
The weights of individual organs were determined and the values were recorded for later quantitative evaluation. Organ samples were digested in 3 ml of concentrated nitric acid (sub-boil grade; produced using a quartz distillation system model MSBQ 2, Maasen, Eningen, Germany) by microwave-assisted digestion. Samples were completely decomposed in pre-cleaned PTFE vessels in a closed, pressurised autoclave system (UniClever, Plazmatronika, Wroclaw, Poland). The digestion programme consisted of three 5-min steps, involving application of 70%, 90% and 100% microwave power (100 W), controlling working pressure within limits of 35/32, 40/35 and 45/42 bar, respectively. After a 10-min cooling period, digests were quantitatively transferred to pre-cleaned PP vials, diluted and adjusted to a final mass of 10 g with ultrapure water (Ultra Clear system, SB Barsbüttel, Germany). Blank samples (typically n = 12 per sampling series) were simultaneously processed in the same way.
Lead content of digests was determined by electrothermal atomic absorption spectrometry (ET AAS), employing AAnalyst 600 Perkin-Elmer (PerkinElmer Inc., Shelton, CT, USA) instrumentation according to the recommended procedure. A mixture of ammonium phosphate and magnesium nitrate was used as a combined chemical modifier. A method of standard addition calibration was applied for quantitation. Lead content of the following organs was determined at the end of week 6 of treatment: lungs, liver, kidney, brain and spleen.
Blood samples were collected by cardiac puncture into 1 ml plastic Eppendorf tubes containing a small amount of heparin. Whole blood was divided into three fractions by centrifugation and proteins were further separated from the remaining supernatant using methanol (300 μl). Samples were stored at 5 °C for subsequent analysis.
Individual fractions of blood were decomposed by means of an ozone and NOx nitrous oxides gas mixture employing the system Dry Mineralizer Apion (Tessek, Prague, Czech Republic) under atmospheric pressure. Blood samples were quantitatively transferred to pre-cleaned glass vials and 1 ml of 1 M nitric acid (high purity sub-boil grade) was added to each sample for enhanced mineralisation. Vials were embedded in a thermo-block heating system in which samples were first dehydrated at 110 °C (ramp time 45 min) for 60 min, then mineralised at 380 °C (ramp time 4 h) for a period 8 h with simultaneous introduction of the gas mixture. After cooling of vials, digest samples were diluted in 1 ml 1 M nitric acid (sub-boil purity), quantitatively transferred to plastic scintillation vials, adjusted with ultrapure water (Ultra Clear system, SB Water Systems, Barsbüttel, Germany) to a final mass of 3 g and stored for subsequent analysis. Blank samples were simultaneously processed in the same manner.
The content of lead in blood digests was determined by electrothermal atomic absorption spectrometry (ETAAS) employing AAnalyst 600 (PerkinElmer Inc., Shelton, CT, USA) instrumentation according to recommended procedures. A mixture of ammonium phosphate and magnesium nitrate was used as a combined chemical modifier. A method of standard addition calibration was applied for quantitation. Lead content was determined at the end of week 6 of exposure.
Results were reported as the mean value ± standard deviation. Data were analysed using Statistica 8.0 software (StatSoft. Inc., Tulsa, OK, USA). Student t-test was used to determine differences between experimental and control groups. Values of p < 0.05 were considered to be statistically significant.
The content of lead in the organ of entry (lung) as well as in the secondary organs was markedly increased after six weeks of nanoparticle inhalation compared to controls (Fig. 1a, b). The highest concentration of lead was found in the kidney, followed by the lungs, liver and spleen, and finally the brain (Fig. 1a, b). The lead concentration in organs of mice exposed to PbO nanoparticles was more than 1500 times higher in lungs, 140 times in kidney, 140 times in liver, 470 times in spleen and about 120 times higher in brain in comparison to the control organs (Additional file 1: Table S1). Similar trend was observed in the second independent experiment (Additional file 1: Table S2).
We also compared lead mass (uptake) after the treatment in all organs that were investigated (Fig. 1c, d). This analysis showed the highest amount of lead to be in the liver, followed by kidney, lungs, and similar lead content was found in spleen and brain.
Moreover, we determined the amount of lead in three fractions of blood (Fig. 2). The largest proportion of lead was found in blood cell fraction (88.2% of lead), lower proportion in the plasma protein fraction (11.6% of lead) and a negligible proportion in serum (below LOD, representing value of about 0.2% of total lead concentration in whole blood). The concentration of lead in whole blood of mice exposed for six weeks to PbO was 132 ng·g−1, whereas this was below the detection limit of 11 ng·g−1 in the control animals.
Histopathological analysis revealed that inhalation of lead oxide nanoparticles caused severe alterations in lung morphology following six weeks exposure. Lungs of these animals exhibited moderate hyperaemia, congested capillaries, enlarged pulmonary septa, haemostasis in basal parts of lobes with presence of many siderophages (macrophages with pigment hemosiderin) and the presence of alveolar emphysema in some areas of the lungs (Fig. 3). Increased numbers of neutrophils and macrophages were found in the alveoli. Bronchioles exhibited the presence of increased mucous secretion, desquamated epithelial cells, neutrophils and sporadic red blood cells. Inhalation of PbO nanoparticles caused also extensive perivascular and peribronchiolar lymphocyte infiltration, which are typical of sub-chronic inflammation. We observed a large number of TUNEL-positive cells in pulmonary basal lobes and in lymphocyte infiltrates (Fig. 4). In addition, elevated numbers of proliferating cells were found in infiltrates, indicating intensive reparative processes initiated in these areas. Activity of Na-K ATPase in lung alveolar tissue was not significantly altered after exposure to PbO nanoparticles (Fig. 4).
Examination using transmission electron microscopy (TEM) demonstrated oedematous enlargement of alveolar epithelial type I (PI) cells and endothelial cells in some regions of the lungs (Fig. 3i-m). Fragmentation of alveolar epithelial type I cells and endothelial cells of the capillaries with denuded basement membranes was also detected. Moreover, the occurrence of apoptotic and necrotic macrophages and granular pneumocytes was observed.
Nanoparticles were located both in alveolar cavities and within cells (Fig. 3i-k'), mainly in alveolar epithelial type I cells (membranous pneumocytes, PI) but not in alveolar epithelial type II cells (granular pneumocytes, PII). Furthermore, they were also found in phagosomes of lung macrophages and neutrophils. In the alveolar cavities, nanoparticles were enveloped in surfactant. Nanoparticles formed clusters within cytoplasmic vesicles. The contents of the vesicles, completely enveloped by cytoplasmic membrane, were more electron-dense than the surrounding cell cytoplasm. Nanoparticles in the vesicles exhibited different sizes, they were well-separated from each other and often located adjacent to the vesicular membrane. Notably, increased amount of tubular myelin (an intermediate step in the formation of surface film) was indicative of enhanced production of surfactant by alveolar epithelial type II cells (PII). Moreover, we observed frequent incidence of thromboses in capillaries and venules (Fig. 3l, m). There were no nanoparticles found in the lungs of control mice exposed to clean air.
Following sub-chronic exposure to PbO nanoparticles, significant changes occurred in liver as a secondary target organ (Fig. 5), mainly manifested by hepatocyte enlargement and hydropic degeneration. Hypetrophy of hepatocytes was combined with karyomegaly, mostly located in centrilobular regions, and with increased amount of binucleated hepatocytes. We found occasional periportal inflammation in some areas, predominantly lymphocytic infiltration. In some samples, areas of hepatic necrosis were observed. PAS staining confirmed impaired glycogen content in hepatocytes (Fig. 5 d, h). There was no evidence of granuloma, noticeable sinusoidal dilatation, hepatic fibrosis (Fig. 5c, g) or bile duct abnormalities.
When examined by TEM, overall morphology of hepatocytes around the central vein was almost same in the animals exposed to nanoparticles as in controls (Fig. 5i-p). Nuclei (1 or 2) were round, small, with smooth contours, with prevalent euchromatin and with few nucleoli. Periportal hepatocytes (zone 1 of liver acinus) instead exhibited typical signs of necrosis. Cells in this area were enlarged with obvious cellular damage, they exhibited clear round nuclei, cytoplasm appeared almost empty and cell organelles were degraded or showed severe pathological signs including hydropic dystrophy (membrane rupture, presence of vacuoles in their bodies and oedema). Periportal hepatocytes are in the most intimate contact with the arterioles, thus representing the best oxygenated hepatocytes and the first group of cells, which absorb blood-borne lead oxide nanoparticles or lead in non-particulate form.
Mitochondria in affected hepatocytes were not evenly distributed throughout the cytoplasm as observed in the controls but they were accumulated around the nucleus and peripherally underneath the plasma membrane. In some cases, they formed short radial septs inside the hepatocytes. Amount of mitochondria exhibiting degeneration of their inner membranes was insignificant. Cisternae of granular endoplasmic reticulum were found in direct contact with mitochondria. In PbO-treated animals, secondary lysosomes (hard upon bile canaliculus) and sporadic peroxisomes were individually dispersed. Lysosomes occurred in the cytoplasm of PbO-treated animals to approximately the same extent as in the controls.
Numerous lipid droplets (tens in a single hepatocyte), resembling electron-dense vacuoles of different sizes, were accumulated in the cytoplasm of hepatocytes (Fig. 5k, l). They either formed groups of lipid droplets or they were located individually with a size ranging from 0.5 to 3 μm. Such accumulation of small intracytoplasmic fat vacuoles (liposomes) is indicative of microvesicular steatosis of liver.
Epithelial cells of the bile ducts were also affected following the exposure to lead oxide nanoparticles. Nuclei and cytoplasm of the lining cells exhibited signs of necrosis. The damage of hepatic endothelial cells was about the same as in bile duct epithelia. In contrast, the ultrastructure of Kupffer cells and Ito cells did not display any changes. Also the numbers of macrophages and Ito cells were unaffected compared to the controls.
Following six weeks of inhalation, nanoparticles were found inside hepatocytes (Fig. 5m-p) where aggregates of nanoparticles accumulated within the centres of their mitochondria. These aggregates were all of about the same size (diameter of about 20–30 nm), they were enveloped by an electron-dense mitochondrial matrix and formed clusters in number of up to tens inside one mitochondrion (Fig. 5m-o). Thus, nanoparticles exhibited different morphology in liver in comparison to the original nanoparticles in the alveolar epithelial cells I. Also their aggregates in livers were smaller and more uniform. Nanoparticles were also found in peroxisomes of hepatocytes possessing the same features as those in mitochondria and they were freely located within the hepatocyte cytoplasm (Fig. 5p). In contrast, nanoparticles were not found in Kupffer cells, Ito cells and bile ducts.
Interestingly, the number of apoptotic cells (TUNEL-positive) in organs of animals exposed to PbO NPs was unchanged in comparison to the controls (Fig. 6). In both, mainly Kupffer cells were TUNEL-positive. Some Kupffer cells also expressed proliferating cell nuclear antigen (PCNA), with their numbers being about the same in PbO-exposed and control animals. In contrast, PCNA-positive hepatocytes (mainly binucleated) were more frequent in Pb-exposed animals compared to the controls. Expression of Na-K ATPase unravealed alterations to functioning of hepatocytes and to sinusoid arrangement. Specifically, activity of Na-K ATPase was almost vanished in some areas of liver lobules following exposure to PbO nanoparticles (Fig. 6).
Chemical composition of livers was analysed by X-EDS to verify lead content of nanoparticles that were visible on ultrathin sections (Fig. 7). Approximately 20 different Region Of Interest (ROI) were analysed. The analysis confirmed the presence of lead in treated samples and no Pb in control samples. Lead was accumulated in the cells and Pb peaks were detected very clearly.
Histopathological analysis revealed that inhalation of lead oxide nanoparticles caused only mild alterations in kidney morphology after sub-chronic exposure (Fig. 8). In some samples, we observed areas with inflammatory infiltrates around renal corpuscles, around renal tubules and blood vessels in the renal cortex. Similar mononuclear cells infiltrates were occasionally found also in control animals. Affected renal corpuscles were slightly enlarged with dilated glomerular capillaries in comparison to the control tissues. The other types of pathological microscopic changes in kidney were rare (Additional file 1 : Table S3, Table S4).
Ultrastructural analysis of kidney tissue unravelled only minimal morphological changes following nanoparticle treatment. Renal tubules showed normal appearance (Fig. 8l), and cortical and medullary collecting ducts had normal structure. Some apoptotic cells were observed in tubules; however, their amount was similar to the controls.
We observed moderate differences in the arrangement of filtration barrier in kidneys of PbO-treated animals. Both laminae rarae were filled by electron-dense material (fine deposits), with lamina densa being thicker compared to the controls. The typical electron contrast among these three layers (laminae) disappeared after lead exposure (Fig. 8k). The filtration membrane was diffusely thickened with intramembranous electron-dense deposits. The distance between the cytoplasmic membrane of endothelial cells and the cytoplasmic membrane of podocytes in the control kidneys was approximately 140–150 nm. Upon exposure to PbO this distance increased to more than 200 nm.
Lead oxide nanoparticles were accumulated in the epithelial cells of the proximal tubules in kidney cortex, the parietal cells of Bowman's capsule and in the epithelial cells of the cortical collecting ducts (Fig. 8m-o). These findings demonstrate the ability of lead oxide nanoparticles to pass through epithelial cells in various parts of the kidney. Lead oxide nanoparticles were dispersed in the cytoplasm but not within cell organelles. No particles were observed inside the nuclei of kidney cells.
Exposure to PbO nanoparticles caused increase in the number of TUNEL-positive cells present in lymphocyte infiltrates (Fig. 9). In other regions of the kidney, TUNEL-positive cells were only rare. In addition, inflammatory infiltrates contained large numbers of proliferating (PCNA-positive) immune cells. Activity of Na-K ATPase was decreased mainly in cells of proximal tubules of animals exposed to PbO nanoparticles.
We did not observe any significant pathological changes in the spleen as a secondary target organ following inhalation of lead oxide nanoparticles (Additional file 1: Figure S5). Animals exposed to PbO nanoparticles exhibited typical spleen morphology undistinguishable from the controls. An arrangement of red pulp and white pulp with blood vessel supply was normal. Areas of erythroblastic islands and regions of granulopoiesis were visible, similar to the controls. The only exception was an increased incidence of megakaryoblasts and megakaryocytes in the pulp of PbO-treated animals (Additional file 1: Figure S5 C, D).
Occurrence of individual lead oxide nanoparticles in spleen sections was rare and these were distributed throughout the cell cytoplasm. Nanoparticles were located in macrophages, erythroblastic cells and reticulocytes (Additional file 1: Figure S5 g, h).
Exposure to lead oxide nanoparticles caused changes (Fig. 10) in the hippocampal region. We observed dark, shrunken, damaged neurons in the pyramidal layer of Ammon's horn, predominantly in CA1 region (Fig. 10d, h). Tracts of nerve fibers surrounding the hippocampal area were also affected. Luxol Fast Blue staining confirmed spongiform changes in these regions (Fig. 10c, d, l, p). Neither the number of PCNA-positive neurons (Additional file 1: Figure S6) nor TUNEL-positive cells was changed after the exposure (data not shown).
In brain tissue, lead oxide nanoparticles were numerous, in contrast to the kidney or spleen (Fig. 10s-u'). The particles were located in cytoplasm of neurons, in their processes and they were also closely associated with synapse regions. Moreover, they were found in pericytes (mural contractile cells of capillaries). Lead oxide nanoparticles were dispersed throughout the cytoplasm or grouped in cytoplasmic vesicles and they exhibited similar appearance to those in the lungs or kidneys.
Currently, the routes of exposure to nanoparticles are numerous and they depend on the conditions related to how and where the nanoparticles are generated and/or employed, e.g. factories, cosmetics, sprays, food, etc. Therefore, same nanoparticles can enter the body by different routes depending on the process of their production. Still, the respiratory tract is the main and typical route by which nanoparticles (NPs) enter the human body [25–27]. This route of entry may continue by multiple secondary distribution avenues, including olfactory path to the brain, translocation to the blood or entry to the GI system during lung clearance.
The lung represents an organ with maximum exposure to PbO nanoparticles and susceptibility to any toxic effects. As macrophages have difficulties identifying such small particles and carrying out phagocytosis, lung defence against nanoparticles is very weak . Nanoparticles are absorbed and deposited in epithelial cells of bronchioles and alveoli, they may rapidly cross the barrier and get absorbed into the circulation, and then migrate to secondary target organs such as the liver and kidney [25, 29]. In agreement with these findings, our analysis revealed increased amount of lead, not just in the primary organ (lung), but also accumulations in the secondary organs. After six weeks of exposure to PbO, the highest concentration of lead was found in the kidney, followed by the lungs, liver, spleen and brain. The same results were previously reported for the treatment with low doses of PbS nanoparticles (kidney > lung > liver) . However, higher doses of PbS nanoparticles resulted in increased amount of lead in the liver compared to the lungs (kidney > liver > lung). Thus, distribution of nanoparticles within the body can be influenced not only by nanoparticle properties but also their dose.
Previous studies revealed large accumulation of lead in the skeleton  at balanced conditions. As we used sub-chronic time point, we can predict that steady condition was not established yet and lead content in bone will not be so high. This is also supported three-compartment model of human lead metabolism postulating that from blood to bone is transported approximately 15% of lead from lead daily intake . Secondary target organs therefore contain proportionally larger amount of total lead in our experimental conditions before their exclusion by urine/feces or their accumulation in skeleton.
However, it is necessary to mention that not all lead found in our target organs is in nanoparticles form. Some nanoparticles can be dissolved after in vivo deposition in the lungs, leading to ions formation, and these ions can be released and transported from the site of their production to other target organs resulting in continual dissolution (and thus ion generation) of the residual nanoparticles . Moreover, the dissolution rate of nanoparticles is not a constant factor but depends on particle size, coating, stability, manufacturing process, and biological environment and unfortunately the literature on nanoparticle dissolution in tissues is sporadic as methodological approaches are recently limited.
The efficient method for the measurement of nanoparticles concentrations in organs is for example inductively coupled plasma mass spectrometry ICP-MS , which was used to evaluate in vivo clearance and translocation of europium-doped gadolinium oxide nanoparticles after 24 h nanoparticle instillation . Analysis revealed 59% of the initial nanoparticle dose in the lung, 20% was excreted in the feces, 0.2% was detected in the liver and less than 0.1% of the delivered dose was found in the remaining organs. These results were qualitatively similar to iridium nanoparticles instilled intratracheally in adult rats after 7 days , when 59% of the delivered dose was found in the lungs, 35% was excreted in the feces and less than 0.2% found in extrapulmonary organs. These in vivo studies confirmed the presence of intratracheally instilled nanoparticles in liver and the other extrapulmonary organs similar as we observed by transmission electron microscope in all analyzed organs and confirmed also by EDX analysis in case of liver. Therefore, the translocation of nanoparticles through the circulation into secondary organs was confirmed but seems to be low , which likely corresponds to our experiments with the majority of nanoparticles observed to stay in the pulmonary tissues. On the other hand, it appears that the accumulation of nanoparticles in target organs might reach a critical threshold causing injury in the case of our sub-chronic exposure as was proposed previously .
Based on analysis the ratio of gadolinium to europium ions measured by ICP-MS in several tissue samples , we can predict amount of particle and non-particle fraction in different organs. The target Gd:Eu ratio was found to be 4.74 based on the initial precursor concentrations of Gd and Eu nitrates . It means that about 4–5 times higher presence of nanoparticles to ions could be present in our tissue samples, therefore pathological changes observed in organs following inhalation of PbO nanoparticles should be associated preferentially with nanoparticle effect and not dissolved form of lead. Unfortunately, similar study was not performed for lead nanoparticles and as different chemical composition of nanoparticles can affect their behavior in vivo, the ratio of Pb nanoparticles and ions can be altered. Ultimate confirmation of proportional lead content in PbO-NP or non-particulate form will be necessary to obtain by ICP-MS analysis in future.
Blood lead level (BLL, μg/dl) is the biologic index used by health care providers as an indicator of lead exposure. Subclinical lead toxicity remains a problem for both adults and children. In 2012, the Centers for Disease Control and Prevention (CDC) adopted 5 μg/dL as the upper reference level for BLLs in children, representing an advisory level for environmental and educational intervention. In 2015, the National Institute for Occupational Safety and Health (NIOSH) designated 5 μg/dL in whole blood from a venous blood sample as the reference blood lead level for adults. An elevated BLL is defined as a BLL ≥ 5 μg/dL . Blood lead levels of 10 μg/dL are associated with aggressive behaviour and hearing dysfunction in children. On the other hand, there is no detectable threshold for adverse effects of lead on the development of the nervous system . After absorption, lead is distributed to the blood, soft tissues and bones. In the blood, 99% of lead is bound to erythrocytes and 1% exists free in plasma, available for exchange with soft tissues (kidney, brain, liver, bone marrow) . In our study following six weeks of exposure by inhalation to lead oxide nanoparticles, we found 88.2% of the lead in blood cell fraction, a lower proportion (11.6%) in the plasma-protein fraction, and only small proportion in the serum (approximately 0.2%). One of possible explanation of this diversity can be caused by the occurrence of lead in multiple forms where dissolved Pb is probably accumulated in blood cell fraction and PbO nanoparticles interact more with proteins in plasma and thus increasing their level in this blood fraction. The concentration of lead in whole blood of mice exposed for six weeks to PbO-NPs was 132 ng/g (with presumptive density of mouse blood 1 g/ml, blood lead level is 13.2 μg/dL), whereas lead concentration was below the detection limit of 11 ng/g in the control animals. The concentration of lead in blood of our exposed mice exceeded 2.6 times a venous blood sample as the reference blood lead level for adults .
Once nanoparticles enter into organs, they penetrate into cells crossing the cytoplasm membrane. Uptake via endocytic routes involves delivery of material into a sub-cellular compartment, the endosome, which is separated from the cell cytoplasm by a membrane. Most endocytic routes end up in degradative compartments of the cell, the lysosomes, where materials are exposed to high concentrations of a wide variety of hydrolytic enzymes. However, there are a few substances, such as heavy metals, which can escape from these recycling activities and accumulate in cells, resulting in toxicity . Our study confirmed such escape of lead oxide nanoparticles from membrane-bounded structures into freely located metal nanoparticles within the cell cytoplasm in all secondary target organs; only in some cases they were located in cytoplasmic vesicles. In the liver, nanoparticles exhibited characteristic arrangement within hepatocyte mitochondria. Previously, the lead has been shown to actively accumulate in the mitochondria . Lead can enter the mitochondria passively at concentrations above 50 μM, inhibiting mitochondrial uptake of Ca2+ and causing displacement of Ca2+. This effect has been demonstrated in isolated mitochondria or in in situ conditions . Lead oxide nanoparticle accumulation inside hepatocytic mitochondria could function like calcium ions. Similarly, prolonged increase in [Ca2+] leads to the opening of the mitochondrial permeability transition pore (PTP), a critical event leading to cell death by apoptosis [39, 40]. Therefore, lead oxide nanoparticles can cause serious damaging events associated with processes occurring just in cell mitochondria.
Inhalation or intra-tracheal instillation of various nanoparticles was previously shown to cause numerous inflammatory changes in lungs [41–44]. Exposure to nanoparticles resulted in serious lung damage, local or systemic inflammation (assessed by bronchoalveolar lavage), increase in macrophage numbers and influx of neutrophils in bronchoalveolar lavage fluid. Comparison of metal oxide nanoparticles, such as CeO2NP, NiONP, ZnONP, and CuONP , revealed a unique inflammatory footprint, both acutely and chronically. Thus, nanoparticles cannot be viewed as a single hazard entity and risk assessment should be performed separately for each substance .
Moreover, metal oxides of nanoparticles can release their soluble ions in vivo during passages via bronchial tree. These ions were shown to cause diverse effect on individual organs in comparison to their nanoparticle forms . Oxides of zinc in aerosol form attracted specifically neutrophils while ZnO in nanoparticle form attracted preferentially eosinophils. Even further, oxides of nickel in aerosol form caused no inflammatory response in contrast to nanoparticle form inducing serious chronic inflammation. On the other hand, oxides of Cu contributed to the lung inflammation attracting neutrophils into lungs both in aerosol form and nanoparticle form. Unfortunately, similar study comparing the effect of lead oxide nanoparticles and lead oxide salts was not performed yet and this comparison will be necessary to follow in future.
Our sub-chronic study with lead oxide nanoparticles confirmed serious pulmonary inflammation after their inhalation, with the presence of siderophages verifying longstanding pulmonary congestion. Moreover, we observed serious thrombotic events in capillaries and small venules in lung tissue caused by lead oxide nanoparticles inhalation. In secondary target organs, no thromboses were found. As lead can cause imbalance of intracellular calcium in tissues and, in addition, nanoparticles may influence blood coagulation [43, 46], this effect of lead oxide nanoparticles is unsurprising. Whether this coagulation is also connected to increased level of calcium in platelets inducing their aggregation in lungs, however, will be necessary to uncover.
In the liver, we observed excess lipid accumulation following lead exposure, indicating impairment of the normal processes of synthesis and elimination of triglycerides. As the liver is the main organ involved in lipid metabolism, it is commonly associated with steatosis. Therefore, the storage disorders (lipid accumulation) and lysosomal perturbations (reduced membrane stability) represent early markers of toxic effects on liver cells and biomarkers of contaminants such as heavy metals . Inhalation of PbO nanoparticles also caused hydropic degeneration or hypetrophy of hepatocytes, focal necrosis and some areas with inflammation, representing typical features of steatohepatitis [48, 49]. It is thought that hydropic degeneration can be caused by hypoxia, ischaemia or the treatment of hepatocytes with endotoxins or other chemicals . This response has also been observed following exposure to other toxic materials, including copper nanoparticles in mice  and cerium oxide nanoparticles in rats .
Surprisingly, morphology of lead oxide nanoparticles was altered in the liver where we found clusters of many small-sized (20–30 nm) agglomerates within hepatocyte mitochondria. Based on this observation, we suggest that disintegration or decomposition of lead oxide nanoparticles occurs preferentially in the liver.
High lead concentration did not cause expected serious changes in kidney. In contrast to lung and liver, we have found only modest changes of kidneys in animals exposed to PbO nanoparticles. Areas of inflammatory infiltrates in the renal cortex were observed, predominantly containing lymphocytes or mononuclear cells. However, such cellular infiltrates are routinely observed in the kidneys of rodents and generally increase with age . Thickened filtration membranes with intramembranous electron-dense deposits were observed, similar to kidney tissue exposed to cadmium nanoparticles . Similarly, rats exposed to silver and gold nanometals demonstrated moderate thickening and a clearly enhanced contour of the glomerular basal membranes .
Lead oxide nanoparticles were found mostly in epithelial cells of proximal tubules and the presence of nanoparticles was consistent with the highest metabolic activity in the kidney occurring in the proximal tubules. By contrast to hepatocytes in liver or alveolar epithelial cells in lung, lead oxide nanoparticles were sparse in these cells. Interestingly, high concentration of lead as shown by chemical analysis did not corresponded to rare presence of lead oxide nanoparticles in kidney cells. Therefore, we propose that the most of kidney lead was present in its dissolved or metabolized form.
Inhalation of lead oxide nanoparticles caused a significant effect on the histological appearance of the hippocampal region in the mouse brain. We demonstrated significantly increased lead content of the brain after lead oxide nanoparticles exposure. These findings are in accordance with studies by others [30, 54] describing pathological changes mainly in the hippocampal regions of nanoparticle treated animals. Lead intoxication in humans causes axonal degeneration and in some other species it causes a primarily demyelinating neuropathy .
Can nanoparticles enter into brain tissues and pass hematoencephalic barrier? After exposure by intranasal instillation, nanoparticles can be deposited in the nasal mucosa and then translocated to the olfactory bulb of the central nervous system (CNS) along the neuronal olfactory route, for example TiO2 nanoparticles  or copper nanoparticles . Here, lead oxide nanoparticles were able to pass hematoencephalic barrier of brain. In our case, we found lead oxide nanoparticles in neurons and in pericyte cytoplasm, indicating blood-brain barrier translocation. We therefore propose this as the main transport route based on the results of our study, however, we cannot exclude transport via the olfactory route as olfactory nerves were not investigated here.
Lead levels increased approximately 120-fold in the brain cortex following six weeks of PbO nanoparticles exposure. Similarly, exposure to PbS nanoparticles  increased lead levels significantly. Lead levels in the cortex were 5.29 times (low dose treatment) and 8.24 times (high dose treatment) higher compared to control samples, and 10.33 times and 13.40 times higher in the hippocampus, respectively. Thus more lead was accumulated in hippocampal area. In this study, male Sprague Dawley rats were exposed to PbS NPs by trachea once every 7 days for 3 consecutive months (15 mg/kg PbS NPs in low exposure group and 30 mg/kg PbS NPs in high exposure group).
It is interesting to note that in our study lead levels in the mouse brain after six weeks of exposure (116 ± 18.13 ng/g) did not reach those in the brain cortex of control animals (0.17 ± 0.04 μg/g) reported by Cao et al. . These discrepancies may be explained by differences between rats and mice, but also by different environmental conditions during both experiments. In spite of this, both studies confirmed increased lead levels in brain cortex after lead nanoparticle exposure.
Lead can cause also imbalances of calcium in nerve tissue . Chronic lead exposure of rats to PbS nanoparticles  increased the [Ca2+]i in nerve cells in comparison to controls. With increased PbS concentration, memory behaviour of animals was influenced, and average number of errors and escape latency were increased. Our study confirmed the presence of lead oxide nanoparticles in brain, therefore, higher attention should be paid to possible negative effects of these inhaled nanoparticles further. They induce serious changes in brain morphology as they can easily penetrate through brain-barriers.
In summary, we found significant microscopic as well as ultramicroscopic changes in primary and secondary organs after lead oxide nanoparticle inhalation. In the lungs, nanoparticles formed clusters inside the cytoplasmic vesicles, while in the liver, kidney, spleen and brain they demonstrated tissue-specific subcellular processing. Our study indicates the variability of nanoparticle transport and intake by individual types of cells in different organs as well as unevenness of lead oxide nanoparticle processing by cells, which will be necessary further address in future to reveal and experimentally enhance the mechanism of their clearance.
Authors would like to thank I. Míšek, D. Klemová, F. Míka, and E. Svozilová for help with the collection and processing of samples.
The work was supported by the Czech Science Foundation (grant no. P503/11/2315 and P503/12/G147) and by the Grant Agency of the Masaryk University (1318/2017).
This manuscript has not been submitted simultaneously for publication in any other journal, nor have the findings been partially disclosed in any other publication.
ZV, MB: conceived and designed experiments, LV, TS, ZV, BD, PM, LC: performed experiments, JD, PF, MB, PC: analyzed the data, AH: contributed reagents and materials, JD, AH, MB, PM: wrote the paper. All authors read and approved the final manuscript.
The experiments were performed in accordance with the ethics policy of the Institute of Animal Physiology and Genetics (no. 081/2010).
The manuscript as submitted has been read and approved by all authors.
Agency for Toxic Substances and Disease Registry (ATSDR). 2016. https://www.atsdr.cdc.gov/csem/csem.asp?csem=7&po=8.


text:
Q: MVC Editar y enviar un Model con View y Partial Views Estoy trabajando en un proyecto con una Vista que tiene un modelo complejo (para simplificar pongo este ejemplo):
public class Person
{
    public string Name { get; set; }
    public string LastName { get; set; }
    public string Data3 { get; set; }
    public string Data4 { get; set; }
    public List<Movie> Movies { get; set; }
}

public class Movie
{
    public string Name { get; set; }
    public double Cost { get; set; }
    public double Quantity { get; set; }
}

Tengo una Vista Principal Persona con dos Vistas Parciales "_Movies" y "_Books".
Cada una de estas vistas parciales tiene un botón "añadir" que abre un popup que solicita los siguientes datos:

*

*Name of the movie

*Cost

*Quantity

La idea es que cuando inserte estos datos, se muestre un grid con la lista de películas y los datos introducidos. Pudiendo editar o borrar estos datos.
Y cuando pulse el botón de la parte inferior "Guardar todo" en la vista principal, se envíen todos los datos del modelo (los de la vista padre y los de las vistas parciales) al controlador.
Pero no se como jugar con los datos del modelo para enviarlos todos juntos.
¿Cómo puedo editar y refrescar los datos de las Vistas Parciales y conseguir enviar al controlador los datos del modelo de la vista principal más los de las vistas parciales?
Adjunto una imagen de ejemplo de lo que estoy intentando.
Por cierto, estoy usando Razor.
¿Podéis darme alguna idea de cómo hacerlo?
No tengo ni idea de cómo abordarlo y eso que a primera vista parece un ejemplo muy simple...
¿Hay alguna forma de no usar javascript/jquery para todo y poder usar el modelo mediante Razor?
Es decir, no me importa usar jQuery para hacer alguna cosa como llamadas ajax o asignar u obtener datos del modelo. Lo que me gustaría saber es si hay alguna forma de tratar con los datos tipados del Modelo desde Razor.
Muchas gracias.

Este sería mi código.
El controlador de la vista principal:
public class PersonController : Controller
{
    // GET: Person
    public ActionResult Index()
    {
        return View();
    }

    public ActionResult Create()
    {
        ViewBag.Movies = new SelectList(new List<SelectListItem> {
            new SelectListItem { Text = "Lord of the rings", Value = "Lord of the rings"},
            new SelectListItem { Text = "Inception", Value = "Inception"}
        }, "Value", "Text");

        return View(new Person());
    }

    [HttpPost]
    public ActionResult Create(Person person)
    {
        try
        {
            // TODO: Add insert logic here

            return RedirectToAction("Index");
        }
        catch
        {
            return View();
        }
    }

    [ChildActionOnly]
    public ActionResult RefreshMovies(List<Movie> movies)
    {
        ViewBag.Movies = new SelectList(new List<SelectListItem> {
            new SelectListItem { Text = "Lord of the rings", Value = "Lord of the rings"},
            new SelectListItem { Text = "Inception", Value = "Inception"}
        }, "Value", "Text");

        return PartialView("~/Views/Person/_Movies.cshtml", movies ?? new List<Movie>());
    }
}

La vista principal:
@model MisPruebasMVC.Models.Person

@{
    ViewBag.Title = "Create";
    Layout = "~/Views/Shared/_Layout.cshtml";
}

General Data

@using (Html.BeginForm())
{
    @Html.AntiForgeryToken()

        @Html.ValidationSummary(true, "", new { @class = "text-danger" })

                @Html.LabelFor(model => model.Name, htmlAttributes: new { @class = "control-label" })
                @Html.EditorFor(model => model.Name, new { htmlAttributes = new { @class = "form-control" } })
                @Html.ValidationMessageFor(model => model.Name, "", new { @class = "text-danger" })

                @Html.LabelFor(model => model.LastName, htmlAttributes: new { @class = "control-label" })
                @Html.EditorFor(model => model.LastName, new { htmlAttributes = new { @class = "form-control" } })
                @Html.ValidationMessageFor(model => model.LastName, "", new { @class = "text-danger" })

                @Html.LabelFor(model => model.Data3, htmlAttributes: new { @class = "control-label" })
                @Html.EditorFor(model => model.Data3, new { htmlAttributes = new { @class = "form-control" } })
                @Html.ValidationMessageFor(model => model.Data3, "", new { @class = "text-danger" })

                @Html.LabelFor(model => model.Data4, htmlAttributes: new { @class = "control-label" })
                @Html.EditorFor(model => model.Data4, new { htmlAttributes = new { @class = "form-control" } })
                @Html.ValidationMessageFor(model => model.Data4, "", new { @class = "text-danger" })

        @Html.Partial("_Movies", Model.Movies ?? new List<MisPruebasMVC.Models.Movie>())

                <input type="submit" value="Create" class="btn btn-default" />

}

La vista parcial _Movies.cshtml
@model IEnumerable<MisPruebasMVC.Models.Movie>

Movies

    <button id="addISPCost" type="button" class="btn btn-primary" data-toggle="modal" data-target="#myModal">
        Add movie
    </button>

New Movie

                <button type="button" class="close" data-dismiss="modal" aria-label="Close">
                </button>

                @{
                    var newMovie = new MisPruebasMVC.Models.Movie();
                }

                    @Html.LabelFor(m => newMovie.Name, "ISP *:", htmlAttributes: new { @class = "control-label col-md-3" })

                        @Html.DropDownList("Id_ISP", new SelectList(ViewBag.Movies, "Value", "Text"), "Select a movie", new { @class = "form-control", @id = "IdISP" })
                        @Html.ValidationMessageFor(m => newMovie.Name, "", new { @class = "text-danger" })

                    @Html.LabelFor(m => newMovie.Cost, "Cost:", htmlAttributes: new { @class = "control-label col-md-3" })

                        @Html.EditorFor(m => newMovie.Cost, new { htmlAttributes = new { @class = "form-control", @id = "Cost" } })
                        @Html.ValidationMessageFor(m => newMovie.Cost, "", new { @class = "text-danger" })

                    @Html.LabelFor(m => newMovie.Quantity, "Quantity:", htmlAttributes: new { @class = "control-label col-md-3" })

                        @Html.EditorFor(m => newMovie.Quantity, new { htmlAttributes = new { @class = "form-control", @id = "MaximoDiario" } })
                        @Html.ValidationMessageFor(m => newMovie.Quantity, "", new { @class = "text-danger" })

                <button type="button" class="btn btn-secondary" data-dismiss="modal">Cerrar</button>
                <button type="button" class="btn btn-primary" id="saveConfiguracionISP">Aceptar</button>

            @Html.DisplayNameFor(model => model.Name)

            @Html.DisplayNameFor(model => model.Cost)

            @Html.DisplayNameFor(model => model.Quantity)

    @foreach (var item in Model)
    {

                @Html.DisplayFor(modelItem => item.Name)

                @Html.DisplayFor(modelItem => item.Cost)

                @Html.DisplayFor(modelItem => item.Quantity)

    }

La vista parcial _Books es igual que la vista parcial _Movies (no lo pongo por simplificar).
He intentado poner las dos vistas parciales en la misma carpeta que la vista padre y llamándolas con:
@Html.Partial("_Movies", Model.Movies ?? new List<MisPruebasMVC.Models.Movie>())

También con:
@Html.Action("RefreshMovies", Model.Movies)

Llamando a esta acción del controlador:
[ChildActionOnly]
public ActionResult RefreshMovies(List<Movie> movies)
{
    ViewBag.Movies = new SelectList(new List<SelectListItem> {
        new SelectListItem { Text = "Lord of the rings", Value = "Lord of the rings"},
        new SelectListItem { Text = "Inception", Value = "Inception"}
    }, "Value", "Text");

    return PartialView("~/Views/Person/_Movies.cshtml", movies ?? new List<Movie>());
}

E incluso las he movido a la carpeta EditorTemplates dentro de Shared llamándolas mediante:
@Html.EditorFor(m=> m.Movies, "_Movies")
@Html.EditorFor(m=> m.Books, "_Books")

Pero no sé cómo añadir las listas de películas y libros de las vistas parciales que tengo en JSON al modelo de la vista padre para enviarlas al controlador en post del BeginForm.
¿Alguna idea?

A: He conseguido enviar al controlador los datos de las vistas en las propiedades Movies y Books.
Dado que por lo que he visto, a la vista sólo le llega el modelo cuando recibe los datos del controlador y luego eso se interpreta como HTML y que sólo se envían al controlador los datos del formulario, lo he resuelto de la siguiente manera:
A la entidad Person, le he añadido las propiedades MoviesRequest y BooksRequest de tipo string, porque aquí me llegarán dos arrays en formato JSON que parsearé a las propiedades Movies y Books respectivamente quedando la clase Person de la siguiente manera:
public class Person
{
    public string Name { get; set; }
    public string LastName { get; set; }
    public string Data3 { get; set; }
    public string Data4 { get; set; }
    public List<Movie> Movies { get; set; }
    public List<Book> Books { get; set; }
    public string MoviesRequest { get; set; }
    public string BooksRequest { get; set; }
}

Por otra parte, en la vista padre he añadido dos campos ocultos que hacen referencia a esas dos propiedades nuevas:
@Html.HiddenFor(m=> m.MoviesRequest, new { id = "hiddenMovies" })
@Html.HiddenFor(m=> m.BooksRequest, new { id = "hiddenBooks" })

En la vista parcial _Books tengo en javascript una variable global libros, la cual en el document ready establezco de la siguiente manera:
libros = @Html.Raw(Json.Encode(Model));

Y cada vez que modifico el array de libros, actualizo el campo hidden, asignándole:
$("#hiddenBooks").val(JSON.stringify(libros));

Lo mismo para la vista parcial _Movies.
Y luego, en el post del formulario, cuando la llamada llega al controlador, lo que hago es parsearlo a su propiedad original de esta forma antes de guardarlo en BD:
[HttpPost]
public ActionResult Create(Person person)
{
    try
    {
        person.Books = JsonConvert.DeserializeObject<List<Books>>(person.BooksRequest);
        person.Movies = JsonConvert.DeserializeObject<List<Movies>>(person.MoviesRequest);

        // TODO: Add insert logic here

        return RedirectToAction("Index");
    }
    catch
    {
        return View();
    }
}

Con esto, puedo editar en cliente los datos como quiera y en el controlador los tengo fuertemente tipados.

redpajama_set_name: RedPajamaStackExchange


Shattered by You is book 3 of the Tear Asunder series by Nashoda Rose; I read this book as a stand-alone. This book focuses on the Crisis, the bass player in Tear Asunder and Haven, the twin sister to Ream who is also in the band (he had his own book also). Haven has been reunited with her twin brother after living through years of horrible, unimaginable situations. She has been living with her brother and his band mates at the farm. Haven uses running to clear her mind when she has become over-whelmed. When she is caught unwisely running during a thunderstorm, Crisis promises not to tell her brother (she doesn't want to cause him any more problems/heartache than she already had) on one condition; when the band goes out on tour, he checks in with her at least once a day to see how she is and how she is dealing with memories of her past. Crisis and Haven soon develop a friendship that begins to develop into deeper feelings for one another but before much can happen between the two of them, Haven must reveal all the hurt that she is bottling up inside her.
Although I have never read any of the other books, I really enjoyed reading Shattered by You. The story flowed and moved along quickly and I found myself not wanting to put the book down. It was nice to see the relationship between Crisis and Haven develop beginning with friendship and leading to more. I enjoyed the banter between these characters. I also thought Crisis' mom stole the spotlight when she was in a scene. If you enjoy rocker books or books about bands, this is a definite read.


With the growing awareness and information regarding preventive measures for combating fatal disease like diabetes, consumers are opting for proper food along with medicines. One can improve the health in a big way by making small changes to the diet, while still enjoying the favorite foods and taking pleasure from the meals. A diabetes diet is simply a healthy eating plan that is high in nutrients, low in fat, and moderate in calories. Manufacturers are now keen towards Diabetes Nutrition on introducing new low calorie food products with sugar substitutes and less oil, in view of the increasing consumer interest toward healthy eating and help prevent diabetes and its concomitant risk factors.


text:
The Biden regime's politicized 'Justice' Department has obviously been given a mandate by the deep state running the current administration: Go after Trump supporters with a vengeance, even if it means lying, cheating, hiding evidence, and creating a wave of false narratives regarding the Jan. 6 'riot' that was actually a provable false flag event.
According to The Epoch Times, at the DOJ's request, a federal court is keeping under wraps an FBI affidavit that completely undermines the regime's seditious conspiracy case against the patriot group Oath Keepers, which shows that members "are not guilty" and "proves that the prosecution is lying to the jury," said an attorney for members of the group last week.
Jonathon Moseley, who previously represented Oath Keepers defendant Kelly Meggs before he had his license to practice law revoked, told the outlet that an FBI interview with a Capitol Police officer proves that in actuality, the Oath Keepers protected the officer from angry rioters near the Capitol Building Rotunda on Jan. 6, 2021.
"This document—together with a photograph of the moment inside the U.S. Capitol on January 6—proves that the prosecution is lying to the jury," Moseley said in the statement. "No one who engages in seditious conspiracy or insurrection stops to come to the aid of the police against the mob.
"If the Oath Keepers were involved in any way in any insurrection or conspiracy to obstruct the Joint Session of Congress on January 6, would they turn and stand between the U.S. Capitol Police against the mob?" Moseley noted. "This is not merely a good act. This is absolute proof that there never was any insurrection or seditious conspiracy. The prosecution's entire case is a fraud upon the American people."
The Epoch Times notes further:
The FBI document in question, a Form 302 summary of Capitol Police officer Harry Dunn's interview with two special agents, came up in open court on Oct. 6 during the seditious-conspiracy trial of Oath Keepers founder Elmer Stewart Rhodes III and four co-defendants, Kelly Meggs, Kenneth Harrelson, Jessica Watkins, and Thomas Caldwell.
Prosecutor Jeffrey Nestler informed Judge Amit Mehta that Moseley was "threatening" to release a sealed document. In response, Mehta took the unusual step of telling the news media covering the trial to post messages on social media to Moseley saying that he would be jailed if he released the document.
Judge Mehta says that if Mosely does he will be in contempt of court and he urges us (reporters) to tweet that out so Moseley will know that. Moseley claims the doc lost its protective order status; Mehta: i decide when something's lost that status. /59
— Roger Parloff (@rparloff) October 6, 2022
"These public tweets from journalists reporting on Mehta's public warning are visible as an incredible event in the already farcical prosecution," Moseley said.
The two FBI agents who interviewed the Officer Dunn on May 18, 2021, asked about the incident in the rotunda. According to video footage taken by an independent journalist, as well as portions of Dunn's interview with the agents that The Epoch Times obtained, "a group of Oath Keepers came upon a volatile standoff in the Small House Rotunda at 2:48 p.m. on Jan. 6," the outlet reported. Dunn and at least two other agitated protesters were shouting at each other, the report added.
It's patently clear that this was not "seditious conspiracy," but rather a concerted effort by the deep state to jam up then-President Donald Trump and demonize his supporters as 'domestic terrorists,' as noted by a trained observer who was at the Capitol Building that day.
J. Michael Waller, who was at the Capitol that day, wrote about what he saw in a piece for The Federalist a week after the breach:
Plainclothes militants. Militant, aggressive men in Donald Trump and MAGA gear at a front police line at the base of the temporary presidential inaugural platform;
Agents-provocateurs. Scattered groups of men exhorting the marchers to gather closely and tightly toward the center of the outside of the Capitol building and prevent them from leaving;
Fake Trump protesters. A few young men wearing Trump or MAGA hats backwards and who did not fit in with the rest of the crowd in terms of their actions and demeanor, whom I presumed to be Antifa or other leftist agitators; and
Disciplined, uniformed column of attackers. A column of organized, disciplined men, wearing similar but not identical camouflage uniforms and black gear, some with helmets and GoPro cameras or wearing subdued Punisher skull patches.
Tagged Under: Biden regime, Capitol Police, conspiracy, deception, deep state, false narrative, false-flag, FBI, FBI affidavit, federal court, intolerance, Jan. 6, Joe Biden, Justice Department, left cult, lies, Oath Keepers, propaganda, White House

redpajama_set_name: RedPajamaCommonCrawl


On-site observations used to be carried out with written notes and photographs. Arup engaged Fusion to develop Arup Inspect, an iPad app enabling easy and accurate data capture onsite. People can upload CAD or working PDF documents prior to inspections and use the app on site to take photos, make notes, and record defects, safely with just one device.
Using GPS locators for each observation, data is pin-pointed to ensure accuracy and eliminate errors. Reports can be previewed, filtered, emailed and additional details added off-site. The time to conduct and compose an audit inspection has been reduced significantly and has become an enjoyable process.
Arup Inspect has been adopted globally, changing the way asset inspections are delivered. Fusion continues to develop specialist versions of the app, providing options to complete structural, fire-safety, seismic and other niche inspection types.


St Catherine's have asked us to sing in County Mall to raise money for their great cause. Come and see us while you are shopping!
A first for Songbirds - a Christmas concert of carols, Christmas songs, audience participation, mince pies and mulled wine. All in aid of our three chosen charities for 2017-2018.
Please come along and join us! Free entry, donations on exit.
We have been asked again this year by Age UK to sing for their charity at the Carfax Bandstand in Horsham.
Come along to the Community Day at St Mary's Church in Southgate, where we will be performing to open the festivities at 11am.
Once again, we will return to Smallbrook Care Home to sing to the residents (including Jo's Grandma!).


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Q: mdns not wait for change but refresh list regularly This code below is running all the time and when there is a change in mdns devices, it prints data.
But I need completely other behaviour. I need to get list of all mdns devices with their data every 100 seconds (so not to wait for change but get whole list at once, then sleep for 100 seconds, and get list again, and again, and again, ...).
How should I edit my code for this behaviour please?
#!/usr/bin/env python
from __future__ import absolute_import, division, print_function, unicode_literals

""" Example of browsing for a service (in this case, HTTP) """

import logging
import socket
import sys
from time import sleep

from zeroconf import ServiceBrowser, ServiceStateChange, Zeroconf

def on_service_state_change(zeroconf, service_type, name, state_change):
    print("Service %s of type %s state changed: %s" % (name, service_type, state_change))

    if state_change is ServiceStateChange.Added:
        info = zeroconf.get_service_info(service_type, name)
        if info:
            print("  Address: %s:%d" % (socket.inet_ntoa(info.address), info.port))
            print("  Weight: %d, priority: %d" % (info.weight, info.priority))
            print("  Server: %s" % (info.server,))
            if info.properties:
                print("  Properties are:")
                for key, value in info.properties.items():
                    print("    %s: %s" % (key, value))
            else:
                print("  No properties")
        else:
            print("  No info")
        print('\n')

if __name__ == '__main__':
    logging.basicConfig(level=logging.DEBUG)
    if len(sys.argv) > 1:
        assert sys.argv[1:] == ['--debug']
        logging.getLogger('zeroconf').setLevel(logging.DEBUG)

    zeroconf = Zeroconf()
    print("\nBrowsing services, press Ctrl-C to exit...\n")
    browser = ServiceBrowser(zeroconf, "_http._tcp.local.", handlers=[on_service_state_change])

    try:
        while True:
            sleep(0.1)
    except KeyboardInterrupt:
        pass
    finally:
        zeroconf.close()

A: I found solution:
(but I don't like one second sleep function at the bottom - it could cut script runtime and show incomplete list)
#!/usr/bin/env python
from __future__ import absolute_import, division, print_function, unicode_literals

""" Example of browsing for a service (in this case, HTTP) """

import logging
import socket
import sys
from time import sleep
import concurrency

from zeroconf import ServiceBrowser, ServiceStateChange, Zeroconf

def zeroconf_info():
    """zeroconf_info returns a list of tuples of the information about other
    zeroconf services on the local network. It does this by creating a
    zeroconf.ServiceBrowser and spending 0.25 seconds querying the network for
    other services."""
    ret_info = []

    def on_change(zeroconf, service_type, name, state_change):
        if state_change is ServiceStateChange.Added:
            info = zeroconf.get_service_info(service_type, name)
            if info:
                address = "{}".format(socket.inet_ntoa(info.address))
                properties = str(info.properties.items())
                item = (str(info.server), address, info.port, properties)
                ret_info.append(item)

    zc = Zeroconf()
    browser = ServiceBrowser(zc, "_http._tcp.local.", handlers=[on_change])
    sleep(1)
    concurrency.concurrent(lambda: zc.close())()
    return ret_info

print (zeroconf_info())


The first thing I wanted to do when DD came under attack was find some sort of camo to wear. A headband, scarf or just pin a camo ribbon on my blouse. My grand kids loved the idea. DD Because of the show, would be easy to show support for. It shouldn't be too hard to come up with ways to outwardly show support.


Kevin Costner plays a death row inmate who is given the powers and thoughts of a deceased CIA agent (played by Ryan Reynolds) in order to finish an important mission. It goes about as well as you'd expect, and soon CIA boss Quaker Wells (Gary Oldman) is on Costner's tail.
This documentary charts the rise of Swedish footballer, karate expert and walking ego Zlatan Ibrahimovic from his difficult upbringing to his outrageous goal-scoring antics across the continent. A must for football fans who want to know the Premier League's new star striker a little better.
Jesse Eisenberg plays a son who returns to his family home after the death of his mother, photographer Isabelle Reid (played by Isabelle Huppert). Once there, he struggles to reconnect with his father (Gabriel Byrne) and his brother (Devin Druid) in the absence of their mother, who they all remember quite differently.
Lazer Team is a sci-fi comedy from the hugely popular internet comedy team Rooster Teeth, best known for web series Red vs Blue. After shooting down a rocket containing a super powered suit, it's up to 4 small town idiots to fend off an alien invasion of Earth.
Dev Patel plays Srinivasa Ramanujan, an Indian mathematician accepted into Cambridge University during World War I, in this biographical drama. Unfortunately, he faces racism and prejudice once he arrives. Jeremy Irons plays Ramanujan's mentor, Professor G.H. Hardy.
In this 8 part adaptation of Stephen King's novel, James Franco plays a school teacher who discovers a time portal that leads to 1960. Franco decides to try and prevent the assassination of JFK, but soon finds that time is against him.
Keep the kids quiet with the latest hit series from everyone's favourite high-pitched critters. In this collection, Alvin, Simon and Theodore are causing trouble at school (although not the kind that would get you expelled, fortunately).
We're off to Norway now for a slice of natural disaster drama. Kristoffer Joner plays a geologist who has just ten minutes to evacuate his family from their sleepy village after a huge landslide triggers an 80 metre tsunami.
In this classic Czech WWII drama, a carpenter is appointed 'Aryan Controller' of a shop owned by a near-blind and deaf Jewish woman unaware of the war. It rightfully won the 1965 Best Foreign Film Oscar.
Josh Duhamel plays Ben, a lawyer determined to bring down the CEO of a corrupt pharmaceutical firm (played by Anthony Hopkins) and strikes gold when the CEO's wife leaks internal emails. However, a mysterious figure on a motorbike soon starts following Ben and his wife… Al Pacino co-stars.
Studio Ghibli's classic movie receives an English overdub features the talents of Daisy Ridley and Dev Patel. Ridley voices Taeko, a young woman who goes on holiday to escape her humdrum life in Tokyo and ends up facing some big decisions about her future happiness.
This popular Chinese series follows a 17 year old Emperor's concubine who has to fight fear, loneliness and politics to survive life in the palace and win the trust of the Emperor.
Empire's Terrence Howard plays Detective Starks, who is investigating a series of copycat killings at the home of imprisoned serial killer Henry Lee Bishop (Peter Fonda). After discovering it plays host to a website called House of Bodies, Starks is forced to ask Bishop for help.
Daryl Hannah star in this sea-based horror, in which a school of sharks start attacking a coastal town after pumping up on chemicals released by a local businessman who wants to transform the town into a glamorous resort.
Love film and TV? Check out 1000s more DVDs and Blu-Rays from just £1.19 on the musicMagpie Store now!


Put on pounds . but naught to ward off man from creating key events that takes us a stride closer to reducing is going to be of work one always be do. With the swells throughout the year that inventions and creations are creating today, day-to-day activities safely quote the iconic American Business magnate Bill gates in saying, "We're reforming the world with technology".
Add our own advancements over technology to assist you to one of most the speediest growing savoir-faire of today, Medical Transcription, and our company have a fabulous potful from overflowing capability which could certainly unclog the blocks while in the console that effectively cause a fabulous snag operating in efficiency. You see, the aim is really to bring in things while automated such as possible, bringing down the available free time users buy figuring on a programs and mouse pointer the computer umpteen periods! The healthcare market is a single one which does have no peacefulness .. Doctors are onto call — leaving all of them with no longer any available free time to carried out their documentation, which is actually also good important concept of patientcare.
SpectraMedi, one particular US found Medical Transcribing company brings come all the way up with deserving implementations within the valuable in advances into Straight forward Flow, the company's workflow oversight system. Chances are they'll have correctly incorporated range features on the road to Easy Edit, one using the Relatively easy Flow web template modules and the actual software designed for online cropping and editing and bringing up-to-date of transcribed documents. So what IS Comfortable EDIT Say files normally created a practitioners dictating the papers into a definite recording phone which 's subsequently wired to Surgery Transcription organizations to seem converted within text framework. There are a short time when your current recordings are able to have track record noise not to mention disturbances setting up the sound recording unclear.
In such a circumstances, Surgical Transcriptionists have always been advised in order to really leave greeting card blanks and hang on further clarification, as over making that guess because to precisely the doubtful words may be. Accuracy is usually of ultimate importance. Virtually error should put these doctor as well as the the course of action into virtually any fiscal responsibility. Such occurrences might be few and furthermore far between, thereby getting it laborious for that doctors with scan into every folder and pick up on files due to blanks. Doing Benistar is even Easy Manage comes in to play. This item identifies and thus filters computer data files containing greeting card blanks into a trustworthy separate folder, making the device easier in support of the chiropractor to visit and manage these data.


Market volatility has been historically low of late. But that's looking more and more like a mirage. When it changes course, the reversal could be explosive.
For the third time in less than a year, a portfolio stock has received a lucrative buyout offer. The board of directors of chemical company Chemtura (CHMT) unanimously agreed to accept a buyout offer from Lanxess AG in a deal valued at $2.5 billion.
Buying call options in Twitter (TWTR) has huge profit potential. With luck, it could be as profitable as my bullish call on Bank of America (BAC) in February - which netted at least one of my readers a $300,000 profit!


For your friend who is allergic to pastels (like me–and fyi the shawl/bracelet/bag come in black) and prefers clean lines even when it comes to diamonds.
Because polka-dots can be minimalist– Yayoi Kusama Festival of Life book, $70.
Le Labo Santal 26 candle, $36. If you could can modernism, I think this is what it would smell like.
Graphic prints that work on the body or hanging on a wall. Hermès Plume H Vibration shawl (which comes in black/white and black/navy), $880.
Who said simple dressers don't like diamonds? Sometimes, lots of them. Cartier Etincelle de Cartier diamond ring, $10,300.
A visit to the former home of the master. Donald Judd NYC home tour tickets, $24.
Lutz Morris belt bag, $850. She might carry less, but it should still be done in style.
Clean, but kind of rock-n-roll. Hermès Medor bracelet, $790.
I love that Hermes shawl in black and navy – one of my favorite combos….


Martins, T.; Midão,L.; Martínez Veiga, S.; Dequech, L; Busse, G.; Bertram, M.; McDonald, A.; Gilliland, G.; Orte, C.; Vives, M. & Costa, E., "Intergenerational programs review: Study design, characteristics of intervention, outcomes and effectiveness", "Journal of Intergenerational Relationships", Volumen 17, Número 1, 2019. Artículo en prensa.


text:
Q: Eclipse opens some .c files incorrectly I'm using a version of Eclipse that came zipped in a folder so I didn't have to install it, it's running in Ubuntu and the project in question is in C and was not created by me.
Most files open correctly, but a few of them are opened in a window with two columns of line numbers and slightly different syntax colours than normal *.c files (here's a screen-shot http://imgur.com/qgd0N). The icon visible in the tab is also different (looks like a text file icon).
Do you have any idea what is causing this?


This laboratory-tested, gentle cleanser stops dandruff while controlling scalp dermatitis and mild psoriasis. Featuring our unique combination of FDA compliant dandruff-control ingredients plus nourishing essential oils of Rosemary, Olive and Jojoba, this medicated cleanser soothes, heals and moisturizes while it eliminates flakes and itchiness. MSM (methylsulfonylmethane) rebalances and conditions the scalp to prevent future dandruff.
Controls recurrence of flaking, scaling and itching associated with dandruff.
Frostified with MSM, Olive, Rosemary Oils & Colloidal Sulfur.


text:
Melanie Martinez's sophomore album, K-12, is another ambitious triumph for the artist. As with her debut album Cry Baby, K-12's music is a vibrant and singular melting pot of low-key hip-hop, soulful pop and indie-leaning electro. K-12's universe is an expansion of the one introduced in Cry Baby. Using lyrics rich with metaphor, songs address the struggle to find a place to belong —including within friendships, the physical world and romantically — even when fitting into society feels like an uphill battle.
Label: ATLANTIC
K-12 [CD/DVD]
Artist: Melanie Martinez
CD - CD
1. Wheels On The Bus
2. Class Fight
3. The Principal
4. Show & Tell
5. Nurse's Office
6. Drama Club
7. Strawberry Shortcake
8. Lunchbox Friends
9. Orange Juice
10. Detention
11. Teacher's Pet
12. High School Sweethearts
13. Recess


In most cases, you can start physical therapy and get the relief you need right now without a physician prescription. You do not need a referral from a physician to begin therapy at Capital Area PT. You have "direct access" to the doctors of physical therapy for 10 visits or 30 days, which can get you going on the right path faster and with fewer appointments. Direct access patients are still covered by most insurance companies (Workers Comp/No Fault and Medicare require a physician prescription).
The therapists are always happy to speak with and include your primary care physician or physician of your choice to coordinate your care at any time. The therapists are also well equipped to determine if a referral to a physician is necessary to complete your care.


text:
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text:
Cuba: Bay of Pigs
Civil Rights: James Meredith
Civil Rights: Birmingham
JFK Assassination (Pulitzer)
China's Atomic Bomb
Vietnam War: Gulf of Tonkin
Civil Rights: Selma
Vietnam War: Tet Offensive
Soviet Invasion of Czechoslovakia
Civil Rights: MLK jr. Assassination
RFK Assassination
Home / Archives
U.S. sympathetic but won't help rebels, says Rusk
WASHINGTON, April 17, 1961 (UPI) -- Secretary of State Dean Rusk said today the anti-Castro invasion of Cuba was not staged from American soil but that the United States is sympathetic with the aims of those participating. Rusk said the Cuban affair was one for the Cubans themselves to settle but that the United States was not indifferent to the extension of Communist tyranny in this hemisphere.
Cuba charges U.S. financed invasion
UNITED NATIONS, N.Y., April 17, 1961 (UPI) -- Cuba charged today that the invasion of Fidel Castro's island was carried out by "mercenaries" coming from Florida and Guatemala and armed and financed by the United States. U.S. Ambassador Adlai Stevenson denied the charge.
Rebel army invades Cuba, battles for beachheads
By FRANCIS L. McCARTHY, UPI Writer
Cuban revolutionaries invaded their island homeland by air and sea today and clashed with Fidel Castro's forces in a violent waterfront battle just 90 miles from Havana.
Invasion crushed, says Castro
By FRANCIS L. McCARTHY
MIAMI, April 20, 1961 (UPI)-Fidel Castro proclaimed triumphantly today that his forces had destroyed an invasion of Cuba and seized a large number of North American-made arms, including heavy Sherman tanks.
Seven including ex-Castroite executed
MIAMI, April 20, 1961 (UPI)-Seven persons, including one identified as a U.S. citizen, were executed by firing squads at dawn in Havana today, Radio Havana announced. It raised the number of executions for the past three days to 24. Radio Havana, in a broadcast monitored here, identified the American citizen as Rafael Diaz Bencom. Among those executed was Humberto Sori Marin, ex-agriculture minister in Premier Fidel Castro's cabinet.
Rebel leader says invasion without U.S. help
NEW YORK, April 21, 1961 (UPI)-Jose Miro Cardona, chief of the exiled anti-Castro forces, declared today that the abortive invasion of Cuba last Monday was mounted without any "military aid" from the United States. "The United States should not intervene militarily against the Castro regime," said Cardona at a press conference here. "I have said repeatedly this is a Cuban fight by Cubans against Cubans."
Cuba reports 400 caught in invasion
MIAMI, April 21, 1961 (UPI) -- Radio Havana today claimed the capture of 400 rebels, including Jose Miro Torra, son of the top Cuban exile leader in the United States. The radio broadcast denounced the captured invaders as "mercenaries" but noted only moments later that some were young men from aristocratic and wealthy Cuban families.
Khrushchev warns U.S. against Cuban invasion
By HENRY SHAPIRO
MOSCOW, April 22, 1961 (UPI) - Premier Nikita S. Khrushchev warned President Kennedy today that an American attack on Cuba might force a Soviet attack against U.S. foreign bases which he said threaten Russia's security.
Kennedy to brief Ike on Cuba
By MERRIMAN SMITH
WASHINGTON, April 22, 1961 (UPI) - President Kennedy planned to meet former President Dwight D. Eisenhower at secluded Camp David, Md., today to give the old soldier a secret report on the crisis in Cuba.
JFK denies U.S. planned Cuba invasion air cover
WASHINGTON, Jan. 24, 1963 (UPI)-President Kennedy said Thursday that the United States never planned to provide air cover for the ill-fated Bay of Pigs invasion of Cuba. He also told a news conference that continuous air surveillance of the island has shown no threatening inflow of arms to the Fidel Castro regime from the Soviet Union.
Next Cuban Missile Crisis
Prev Berlin Wall
More News from 1960-1969


Oscar, Program Director of Chinaccelerator, is a "Full stack Innovator" with expertise ranging from Product Development to corporate strategy. He has experience as an entrepreneur, Management consultant and early stage investor.
Telecom MEng and Biomedical PhD dropout, began his career 14 years ago as an R&D engineer working in Reseach Center for Ericsson and Telefonica and after discovering Business Model Innovation transitioned to the world of startups. After several years working for a MedTech Startup moved to China in early 2008. Since his arrival to China has been involved in Innovation: product launches, go-to market strategies, Venture Capital financing and M&As for startups and multinationals.
SOSV began supplying start-ups with rocket fuel 20 years ago. We are now a global fund with >$250M assets under management and a staff of nearly 50 operating world-renowned accelerators in the areas of hardware, software, biology, food, robotics, medical devices, transportation, green energy, and beyond. We have over 400 companies in our portfolio, and graduate over 150 start-ups from our accelerator programs each year., Innovation and Venture Capital.