Remaining microtia tissue as a source for 3D bioprinted elastic cartilage tissue constructs, potential use for surgical microtia reconstruction.

The absence of ears in children is a global problem. An implant made of costal cartilage is the standard procedure for ear reconstruction; however, side effects such as pneumothorax, loss of thoracic cage shape, and respiratory complications have been documented. Three-dimensional (3D) printing allows the generation of biocompatible scaffolds that mimic the shape, mechanical strength, and architecture of the native extracellular matrix necessary to promote new elastic cartilage formation. We report the potential use of a 3D-bioprinted poly-ε-caprolactone (3D-PCL) auricle-shaped framework seeded with remaining human microtia chondrocytes for the development of elastic cartilage for autologous microtia ear reconstruction. An in vivo assay of the neo-tissue formed revealed the generation of a 3D pinna-shaped neo-tissue, and confirmed the formation of elastic cartilage by the presence of type II collagen and elastin with histological features and a protein composition consistent with normal elastic cartilage. According to our results, a combination of 3D-PCL auricle frameworks and autologous microtia remnant tissue generates a suitable pinna structure for autologous ear reconstruction.

In Vitro Evidence of Differential Immunoregulatory Response between MDA-MB-231 and BT-474 Breast Cancer Cells Induced by Bone Marrow-Derived Mesenchymal Stromal Cells Conditioned Medium.

Inside tumors, cancer cells display several mechanisms to create an immunosuppressive environment. On the other hand, by migration processes, mesenchymal stromal cells (MSCs) can be recruited by different cancer tumor types from tissues as distant as bone marrow and contribute to tumor pathogenesis. However, the impact of the immunoregulatory role of MSCs associated with the aggressiveness of breast cancer cells by soluble molecules has not been fully elucidated. Therefore, this in vitro work aimed to study the effect of the conditioned medium of human bone marrow-derived-MSCs (hBM-MSC-cm) on the immunoregulatory capability of MDA-MB-231 and BT-474 breast cancer cells. The hBM-MSC-cm on MDA-MB-231 cells induced the overexpression of TGF-ß, IDO, and IL-10 genes. Additionally, immunoregulation assays of mononuclear cells (MNCs) in co-culture with MDA-MB-231 and hBM-MSC-cm decreased lymphocyte proliferation, and increased proteins IL-10, TGF-ß, and IDO while also reducing TNF levels, shooting the proportion of regulatory T cells. Conversely, the hBM-MSC-cm did not affect the immunomodulatory capacity of BT-474 cells. Thus, a differential immunoregulatory effect was observed between both representative breast cancer cell lines from different origins. Thus, understanding the immune response in a broader tumor context could help to design therapeutic strategies based on the aggressive behavior of tumor cells.

Geospatial analysis as a tool to identify target areas for Chagas disease education for healthcare providers.

Chagas Disease (CD) is a neglected zoonotic disease of the Americas. It can be fatal if not diagnosed and treated in its early stages. Using geospatial and sensitivity analysis, this study focuses on understanding how to better allocate resources and educational information to areas in the United States, specifically Texas, that have the potential for increased risk of CD cases and the associated costs of addressing the disease. ICD-9 and 10 inpatient hospital diagnostic codes were used to illustrate the salience of potentially missed CD diagnoses (e.g., cardiomyopathic diagnoses) and where these are occurring with more frequency. Coding software along with GIS and Microsoft Excel 3D mapping were used to generate maps to illustrate where there may be a need for increased statewide surveillance and screening of populations at greater risk for CD. The CD cases reported to the Texas Department of State Healthcare Services (TxDSHS) are not homogenously dispersed throughout the state but rather, reveal that the incidences are in clusters and primarily in urban areas, where there is increased access to physician care, CD research and diagnostic capabilities.

Effect of Moringa oleifera seed extract on antimicrobial activity and in vitro fertilization ability of cryopreserved ram semen.

Cryopreservation has adverse effects on the post-thaw sperm quality due to oxidative stress and the presence of bacteria. To minimize such effects, plant extracts have been included in the composition of the semen diluents. The objective of the present study was to evaluate the antimicrobial and antioxidant effects of Moringa oleifera seed extract (MOSE) on cryopreserved ram semen, as well as its impact on in vitro fertilization. Semen from six hair rams was treated with five treatments before cryopreservation: Control (without any antibiotic), Standard (conventional antibiotic), 1.0, 10.0, and 50.0 mg/ml of MOSE. Post-thawing sperm characteristics were evaluated by the computer-assisted semen analysis. Antimicrobial activity was assessed by counting colony-forming units (CFU) and the antioxidant capacity by the ferric reducing antioxidant power method. A heterologous in vitro fertilization technique was implemented to measure the fertilization rate. Progressive and rapid motility, membrane and acrosome integrity, and active mitochondria were higher (p < .05) in the 10.0 mg/ml treatment compared with Standard after thawing. All M. oleifera treatments showed inhibition of CFU. The antioxidant capacity of M. oleifera seed extract was higher in the 10.0 and 50.0 mg/ml treatments. Fertilization rate (cleavage percentage) was higher (p < .05) in the 10.0 mg/ml (82.9 ± 10.0) and Control (82.5 ± 9.9) treatments compared with Standard (73.7 ± 9.1). The addition of 10.0 mg/ml of MOSE to ram semen inhibits the development of microorganisms and improves sperm characteristics and the in vitro fertility of the semen.

Heparan Sulfate and Sialic Acid in Viral Attachment: Two Sides of the Same Coin?

Sialic acids and heparan sulfates make up the outermost part of the cell membrane and the extracellular matrix. Both structures are characterized by being negatively charged, serving as receptors for various pathogens, and are highly expressed in the respiratory and digestive tracts. Numerous viruses use heparan sulfates as receptors to infect cells; in this group are HSV, HPV, and SARS-CoV-2. Other viruses require the cell to express sialic acids, as is the case in influenza A viruses and adenoviruses. This review aims to present, in a general way, the participation of glycoconjugates in viral entry, and therapeutic strategies focused on inhibiting the interaction between the virus and the glycoconjugates. Interestingly, there are few studies that suggest the participation of both glycoconjugates in the viruses addressed here. Considering the biological redundancy that exists between heparan sulfates and sialic acids, we propose that it is important to jointly evaluate and design strategies that contemplate inhibiting the interactions of both glycoconjugates. This approach will allow identifying new receptors and lead to a deeper understanding of interspecies transmission.

Provision of Palliative Care during the COVID-19 Pandemic: A Systematic Review of Ambulatory Care Organizations in the United States.

Background and objectives: Ambulatory (outpatient) healthcare organizations continue to respond to the COVID-19 global pandemic using an array of initiatives to sustain a continuity of palliative care. Continuance of palliative care during major crises has been previously accomplished; however, the global pandemic presents new challenges to the US healthcare industry. Materials and methods: This systematic review queried four research databases to identify applicable studies related to the provision of palliative care during the pandemic in outpatient organizations within the United States. Results: There are two primary facilitators for the ongoing provision of palliative care for the outpatient segment of the United States healthcare industry: technology and advanced care planning. Researchers also identified two primary barriers in the outpatient setting impacting the continuance of palliative care: lack of resources and accessibility to care. Conclusions: This systematic review identified facilitators and barriers for palliative care initiatives in the United States that can further assist future outpatient (ambulatory care) providers at a global level as the pandemic and associated public health initiatives continue.

Genetic ablation of SLK exacerbates glomerular injury in adriamycin nephrosis in mice.

Ste20-like kinase SLK is critical for embryonic development and may play an important role in wound healing, muscle homeostasis, cell migration, and tumor growth. Mice with podocyte-specific deletion of SLK show albuminuria and damage to podocytes as they age. The present study addressed the role of SLK in glomerular injury. We induced adriamycin nephrosis in 3- to 4-mo-old control and podocyte SLK knockout (KO) mice. Compared with control, SLK deletion exacerbated albuminuria and loss of podocytes, synaptopodin, and podocalyxin. Glomeruli of adriamycin-treated SLK KO mice showed diffuse increases in the matrix and sclerosis as well as collapse of the actin cytoskeleton. SLK can phosphorylate ezrin. The complex of phospho-ezrin, Na+/H+ exchanger regulatory factor 2, and podocalyxin in the apical domain of the podocyte is a key determinant of normal podocyte architecture. Deletion of SLK reduced glomerular ezrin and ezrin phosphorylation in adriamycin nephrosis. Also, deletion of SLK reduced the colocalization of ezrin and podocalyxin in the glomerulus. Cultured glomerular epithelial cells with KO of SLK showed reduced ezrin phosphorylation and podocalyxin expression as well as reduced F-actin. Thus, SLK deletion leads to podocyte injury as mice age and exacerbates injury in adriamycin nephrosis. The mechanism may at least in part involve ezrin phosphorylation as well as disruption of the cytoskeleton and podocyte apical membrane structure.

Assessing the effectiveness of Chagas disease education for healthcare providers in the United States.

BACKGROUND: Chagas disease is a zoonotic infection caused by the parasite Trypanosoma cruzi, which affects an estimated 8-11 million people globally. Chagas disease is almost always associated with poverty in rural areas and disproportionately impacts immigrants from Latin America living in the United States. Approximately 20-30% of people who are infected with Chagas disease will develop a chronic form of the infection that can be fatal if left untreated. Chagas disease is vastly underestimated in the United States, often goes undiagnosed and is not well understood by most U.S. healthcare providers. One of the most important ways at reducing barriers to improving diagnostics of Chagas disease in the U.S. is giving healthcare providers the most up-to-date information and access to leading experts. METHODS: An online webinar was conducted for healthcare providers, veterinarians and public health professionals using Chagas disease expert panelists. Pre and post tests were administered to participants (n = 57) to determine the efficacy in raising awareness and to determine key focus areas for improving knowledge. A Wilcoxon rank-sum was used for non-parametric variables equivalent and for questions that assessed knowledge the McNemar's Chi-Square test was used. RESULTS: There were statistically significant learning increases in multiple categories including transmission (p = <.001), clinical presentation (p = 0.016), diagnostics (p = <.001), and treatment (p = <.001). CONCLUSION: Providing easily accessible learning opportunities using validated testing and evaluations should be further developed for rural healthcare providers in the U.S. as well as healthcare providers serving under represented populations such as immigrants. There is a clear lack of knowledge and awareness surrounding Chagas disease in the United States and just by raising awareness and providing education on the topic, lives will be saved.

Barriers to the Use of Mobile Health in Improving Health Outcomes in Developing Countries: Systematic Review.

BACKGROUND: The use of mobile health (mHealth) technologies to improve population-level health outcomes around the world has surged in the last decade. Research supports the use of mHealth apps to improve health outcomes such as maternal and infant mortality, treatment adherence, immunization rates, and prevention of communicable diseases. However, developing countries face significant barriers to successfully implement, sustain, and expand mHealth initiatives to improve the health of vulnerable populations. OBJECTIVE: We aimed to identify and synthesize barriers to the use of mHealth technologies such as text messaging (short message service [SMS]), calls, and apps to change and, where possible, improve the health behaviors and health outcomes of populations in developing countries. METHODS: We followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses checklist. Deriving search criteria from the review's primary objective, we searched PubMed and CINAHL using an exhaustive terms search (eg, mHealth, text messaging, and developing countries, with their respective Medical Subject Headings) limited by publication date, English language, and full text. At least two authors thoroughly reviewed each article's abstract to verify the articles were germane to our objective. We then applied filters and conducted consensus meetings to confirm that the articles met the study criteria. RESULTS: Review of 2224 studies resulted in a final group of 30 articles for analysis. mHealth initiatives were used extensively worldwide for applications such as maternal health, prenatal care, infant care, HIV/AIDS prevention, treatment adherence, cardiovascular disease, diabetes, and health education. Studies were conducted in several developing countries in Africa, Asia, and Latin America. From each article, we recorded the specific health outcome that was improved, mHealth technology used, and barriers to the successful implementation of the intervention in a developing country. The most prominent health outcomes improved with mHealth were infectious diseases and maternal health, accounting for a combined 20/30 (67%) of the total studies in the analysis. The most frequent mHealth technology used was SMS, accounting for 18/30 (60%) of the studies. We identified 73 individual barriers and grouped them into 14 main categories. The top 3 barrier categories were infrastructure, lack of equipment, and technology gap, which together accounted for 28 individual barriers. CONCLUSIONS: This systematic review shed light on the most prominent health outcomes that can be improved using mHealth technology interventions in developing countries. The barriers identified will provide leaders of future intervention projects a solid foundation for their design, thus increasing the chances for long-term success. We suggest that, to overcome the top three barriers, project leaders who wish to implement mHealth interventions must establish partnerships with local governments and nongovernmental organizations to secure funding, leadership, and the required infrastructure.

Seroprevalence of anti-hepatitis E virus antibodies in domestic pigs in Mexico.

BACKGROUND: Hepatitis E virus (HEV) infection is one of the most common causes of acute liver diseases in humans worldwide. In developing countries, HEV is commonly associated with waterborne outbreaks. Conversely, in industrialized countries, HEV infection is often associated with travel to endemic regions or ingestion of contaminated animal products. Limited information on both, human and animal HEV infection in Mexico is available. As a consequence, the distribution of the virus in the country is largely unknown. Here, we assessed the seroprevalence of HEV among swine in different geographical regions in Mexico. METHODS: Seroprevalence of anti-HEV antibodies in swine herds in Mexico was evaluated in a representative sample including 945 pig serum specimens from different regions of the country using a commercial enzyme-linked immunosorbent assay (ELISA). RESULTS: The overall prevalence of anti-HEV antibodies in swine was 59.4%. The northern region of Mexico exhibited the highest seroprevalence in the country (86.6%), while the central and southern regions in Mexico showed lower seroprevalence, 42.7% and 51.5%, respectively. CONCLUSIONS: In Mexico, HEV seroprevalence in swine is high. Importantly, northern Mexico showed the highest seroprevalence in the country. Thus, further studies are required to identify the risk factors contributing to HEV transmission among pigs in the country. Assessment of HEV human infection in the context of viral transmission in swine is required to better understand the epidemiology of hepatitis E in Mexico.

Bicipital tuberosity bone characteristics in surgical reattachment of the distal biceps: anatomical and radiological study.

PURPOSE: The aim of this study was to measure the cortical thickness and bone density of the different parts of the bicipital tuberosity, to evaluate the importance of these variables on resistance to pulling out of distal biceps tendon reinsertion implants. METHODS: Sixteen cadaveric arms were used for this study. A multiple detector computed tomography was performed in each proximal radius. Bone thickness and density of anterior, posterior cortex and anterior trabecular bone were measured in proximal, medial and distal parts of the bicipital tuberosity. Statistical and concordance analyses of results were performed. RESULTS: In our specimens, the medial and distal parts of the anterior cortex and the anterior trabecular bone were thicker, mean 11.3 mm SD 2.72 and 11.17 mm SD 3.05, with a significant difference when compared to the proximal part; mean 10.3 mm SD 2.35, of radial tuberosity. The three posterior segments where all thicker compared to the anterior cortex (proximal 3.15 SD 1.31; medial 3.33 SD 1.5; distal 3.34 SD 1.43 mm), but without statistical differences between them. The measured bone density was equivalent in the three portions of the anterior cortex and trabecular bone [proximal 1924.63 SD 547.22; medial 1848.19 SD 538.59; distal 2100.47 SD 396.32 Hounsfield units (HU)]. The posterior cortex was denser compared to the anterior cortex and the anterior trabecular bone in all the segments (proximal 1962.63 SD 223.57; medial 1907.16 SD 232.08; distal 1987.06 SD 189.12 HU), but without statistical differences between the three parts. CONCLUSIONS: Based on the results of this anatomic study which have demonstrated that anterior cortex and anterior trabecular bone of the medial and distal regions of the bicipital tuberosity are thicker than proximal part, we postulate that these segments could give better pulling out resistance to monocortical implants. Our findings suggest that the strongest parts of the bicipital tuberosity are the proximal and medial parts of the posterior cortex. We can afford them drilling across the radius using a bicortical implant in the proximal and medial section of the radial tuberosity. Furthermore, we suggest that an increased margin of safety could be achieved to prevent injury to the posterior interosseous nerve, drilling the cortical hole in the proximal part of the radial tuberosity without losing resistance properties.

A life-threatening situation due to a spider bite : a non-infectious necrotizing fasciitis.

We present the case of a healthy patient who sustained a spiderbite in the elbow and developed a non-infectious necrotizing fasciitis in the affected limb. Female patient aged 24 pain reported a spiderbite received some 72 h previously in Mexico (the spider was identified as a brown recluse spider-Loxosceles reclusa). Under the suspected diagnosis of necrotizing fasciitis urgent surgery was indicated. During her hospital stay, the patient required three additional surgical procedures, and was discharged from hospital 30 days after admission. Spider bites in the limb may be limb-threatening and life-threatening. Emergency doctors should be aware of this possibility, because spiders can be unintentionally transported all over the world.

Intradialytic Hypertension / Hypotension and Mortality in San Juan, Puerto Rico.

End Stage Renal Disease patients undergo profound hemodynamic changes during hemodialysis treatments which are now recognized as a marker for increased risk of morbidity and mortality. Development of intradialytic hypotension or hypertension are a common clinical problem in this population with an incidence of up to 20%. We performed a retrospective review of 49 Hispanic patients receiving ambulatory hemodialysis during a period of 6 months to ascertain the development of aforementioned intradialytic events. Clinical data examined the association of these events to mortality and their relationship to antihypertensive medications and cardiomegaly. The prevalence of intradialytic hypotension was 38.78%, hypertension 16.33% individually and both taking place 16.33%. Taken together, the prevalence of these intradialytic events was 71.43% in our Hispanic population. A significant association was found between mortality and Beta blockers (BB)(P=0.044), Calcium channel blockers (CCB) (P=0.023), cardiomegaly (P=0.044), and intradialytic events (P=0.035). Odds ratio of multiple variables dis- closed that dependent variable death decreased in probability with the use of BB by an estimate of 73% and with the use of CCB by 74.8%. On the other hand, odds of developing the dependent variable death increased by 74.5% if the patients developed intradialytic events. Similarly, the odds of developing cardiomegaly in the living group increased by 70%. A logistic regression of multiple variables found that the probability of developing the dependent condition of death increases by almost 2.896 times if intradialytic events are present and that there is a 58.9% inferred causality. It is concluded that intradialytic hyper- tension and hypotension are major risk factors for mortality in dialysis patients. The use of BB and CCB may be protetive to avoid the risk of mortality in these patients.

Tuning thermoresponsive supramolecular G-quadruplexes.

Thermoresponsive systems are attractive due to their suitability for fundamental studies as well as their practical uses in a wide variety of applications. While much progress has been achieved using polymers, alternative strategies such as the use of well-defined nonpolymeric supramolecules are still underdeveloped. Here we report three 8-aryl-2'-deoxyguanosine derivatives (8ArGs) that self-assemble in aqueous media into precise thermoresponsive supramolecular G-quadruplexes (SGQs). We report the synthesis of such derivatives, studies of their isothermal self-assembly, and the thermally induced assembly to form higher-order meso-globular assemblies we term supramolecular hacky sacks (SHS). The lower critical solution temperature (LCST) that indicates the formation of the SHS was modulated by changing (a) intrinsic parameters (i.e., structure of the 8ArGs); (b) extrinsic parameters such as the salt used to promote the formation of the SGQ; and (c) supramolecular parameters such as the coassembly different 8ArGs to form heteromeric SGQs. Changes in the intrinsic parameters lead to LCST variations in the range of 28-59 °C. Modulating extrinsic parameters such as replacing KI with KSCN abolishes the thermoresponsive phenomenon whereas changing the cation from K(+) to Na(+) or adjusting the pH (in the range of 6-8) has negligible effects on the LCST. Modulating supramolecular parameters results in transition temperatures that are intermediate between those obtained by the respective homomeric SGQs, although the specific proportions of the subunits are critical in determining the reversibility of the process. Given the extensive applications of thermoresponsive polymers, the nonpolymeric supramolecular counterparts presented here may represent an attractive alternative for fundamental studies and biorelevant applications.

Interventional nephrology in Puerto Rico: a four year experience.

Puerto Rico is one of the most prevalent areas covered by Medicare in need of renal replacement therapy for which interventional procedures are performed. A cumulative analysis of this management is reported in patients during the period between June 2007 and August 2010. Experience accumulated with 3755 surgical patients revealed that 58% had intravascular catheters, 28% had arteriovenous fistulas, 15% had arteriovenous grafts, and 2% without vascular access. Procedures performed in these patients were: catheter introduction in 1990 cases (33%), angioplasty in 751 cases (20%), angiography in 450 cases (12%), thrombectomy in 413 cases (11%) and venous mapping in 151 cases (4%). The success rates of these procedures were evaluated by analysis of the Society of Interventional Radiology (SIR) criteria for Lifeline Vascular Access. Using SIR definition of success rate for at least one session that includes "declots", placement of catheters and angioplasty, our results revealed an average of 98.2% overall success rate greater than the standard value KDOQI/SIR (> 85% ). This study has documented for four years the success rate of Vascular Interventional Nephrology Center at Auxilio Mutuo Hospital. In order to maintain this success rate is necessary to further evaluate its effectiveness and, most importantly, the development of an educational program for vascular access in patients with chronic kidney disease prior to placement in dialysis units.

The effect of COVID-19 on cancer incidences in the U.S.

Fundamental data analysis assists in the evaluation of critical questions to discern essential facts and elicit formerly invisible evidence. In this article, we provide clarity into a subtle phenomenon observed in cancer incidences throughout the time of the COVID-19 pandemic. We analyzed the cancer incidence data from the American Cancer Society [1]. We partitioned the data into three groups: the pre-COVID-19 years (2017, 2018), during the COVID-19 years (2019, 2020, 2021), and the post-COVID-19 years (2022, 2023). In a novel manner, we applied principal components analysis (PCA), computed the angles between the cancer incidence vectors, and then added lognormal probability concepts in our analysis. Our analytic results revealed that the cancer incidences shifted within each era (pre, during, and post), with a meaningful change in the cancer incidences occurring in 2020, the peak of the COVID-19 era. We defined, computed, and interpreted the exceedance probability for a cancer type to have 1000 incidences in a future year among the breast, cervical, colorectal, uterine corpus, leukemia, lung & bronchus, melanoma, Hodgkin's lymphoma, prostate, and urinary cancers. We also defined, estimated, and illustrated indices for other cancer diagnoses from the vantage point of breast cancer in pre, during, and post-COVID-19 eras. The angle vectors post the COVID-19 were 72% less than pre-pandemic and 28% less than during the pandemic. The movement of cancer vectors was dynamic between these eras, and movement greatly differed by type of cancer. A trend chart of cervical cancer showed statistical anomalies in the years 2019 and 2021. Based on our findings, a few future research directions are pointed out.

Genetic deletion of calcium-independent phospholipase A2γ protects mice from diabetic nephropathy.

Calcium-independent phospholipase A2γ (iPLA2γ) is localized in glomerular epithelial cells (GECs)/podocytes at the mitochondria and endoplasmic reticulum, and can mediate release of arachidonic acid and prostanoids. Global knockout (KO) of iPLA2γ in mice did not cause albuminuria, but resulted in mitochondrial structural abnormalities and enhanced autophagy in podocytes. In acute glomerulonephritis, deletion of iPLA2γ exacerbated albuminuria and podocyte injury. This study addresses the role of iPLA2γ in diabetic nephropathy. Hyperglycemia was induced in male mice with streptozotocin (STZ). STZ induced progressive albuminuria in control mice (over 21 weeks), while albuminuria did not increase in iPLA2γ KO mice, remaining comparable to untreated groups. Despite similar exposure to STZ, the STZ-treated iPLA2γ KO mice developed a lower level of hyperglycemia compared to STZ-treated control. However, there was no significant correlation between the degree of hyperglycemia and albuminuria, and even iPLA2γ KO mice with greatest hyperglycemia did not develop significant albuminuria. Mortality at 21 weeks was greatest in diabetic control mice. Sclerotic glomeruli and enlarged glomerular capillary loops were increased significantly in diabetic control compared to diabetic iPLA2γ KO mice. Glomerular matrix was expanded in diabetic mice, with control exceeding iPLA2γ KO. Glomerular autophagy (increased LC3-II and decreased p62) was enhanced in diabetic iPLA2γ KO mice compared to control. Treatment of cultured GECs with H2O2 resulted in increased cell death in control GECs compared to iPLA2γ KO, and the increase was slightly greater in medium with high glucose compared to low glucose. H2O2-induced cell death was not affected by inhibition of prostanoid production with indomethacin. In conclusion, mice with global deletion of iPLA2γ are protected from developing chronic glomerular injury in diabetic nephropathy. This is associated with increased glomerular autophagy.

Role of the Ste20-like kinase SLK in podocyte adhesion.

SLK controls the cytoskeleton, cell adhesion, and migration. Podocyte-specific deletion of SLK in mice leads to podocyte injury as mice age and exacerbates injury in experimental focal segment glomerulosclerosis (FSGS; adriamycin nephrosis). We hypothesized that adhesion proteins may be substrates of SLK. In adriamycin nephrosis, podocyte ultrastructural injury was exaggerated by SLK deletion. Analysis of a protein kinase phosphorylation site dataset showed that podocyte adhesion proteins-paxillin, vinculin, and talin-1 may be potential SLK substrates. In cultured podocytes, deletion of SLK increased adhesion to collagen. Analysis of paxillin, vinculin, and talin-1 showed that SLK deletion reduced focal adhesion complexes (FACs) containing these proteins mainly in adriamycin-induced injury; there was no change in FAC turnover (focal adhesion kinase Y397 phosphorylation). In podocytes, paxillin S250 showed basal phosphorylation that was slightly enhanced by SLK; however, SLK did not phosphorylate talin-1. In adriamycin nephrosis, SLK deletion did not alter glomerular expression/localization of talin-1 and vinculin, but increased focal adhesion kinase phosphorylation modestly. Therefore, SLK decreases podocyte adhesion, but FAC proteins in podocytes are not major substrates of SLK in health and disease.

Deletion of IRE1α in podocytes exacerbates diabetic nephropathy in mice.

Protein misfolding in the endoplasmic reticulum (ER) of podocytes contributes to the pathogenesis of glomerular diseases. Protein misfolding activates the unfolded protein response (UPR), a compensatory signaling network. We address the role of the UPR and the UPR transducer, inositol-requiring enzyme 1α (IRE1α), in streptozotocin-induced diabetic nephropathy in mice. Diabetes caused progressive albuminuria in control mice that was exacerbated in podocyte-specific IRE1α knockout (KO) mice. Compared to diabetic controls, diabetic IRE1α KO mice showed reductions in podocyte number and synaptopodin. Glomerular ultrastructure was altered only in diabetic IRE1α KO mice; the major changes included widening of podocyte foot processes and glomerular basement membrane. Activation of the UPR and autophagy was evident in diabetic control, but not diabetic IRE1α KO mice. Analysis of human glomerular gene expression in the JuCKD-Glom database demonstrated induction of genes associated with the ER, UPR and autophagy in diabetic nephropathy. Thus, mice with podocyte-specific deletion of IRE1α demonstrate more severe diabetic nephropathy and attenuation of the glomerular UPR and autophagy, implying a protective effect of IRE1α. These results are consistent with data in human diabetic nephropathy and highlight the potential for therapeutically targeting these pathways.

Layered Double Hydroxides (LDH) as Delivery Vehicles of a Chimeric Protein Carrying Epitopes from the Porcine Reproductive and Respiratory Syndrome Virus.

The Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) causes reproductive failure and respiratory symptoms, leading to huge economic losses for the pig farming industry. Although several vaccines against PRRSV are available in the market; they show an overall low efficacy, and several countries have the need for vaccines covering the local, circulating variants. This project aims at developing a new chimeric antigen targeting specific epitopes from PRRSV and evaluating two test adjuvants to formulate a vaccine candidate. The test antigen was called LTB-PRRSV, which was produced recombinantly in Escherichia coli and consisted of the heat labile enterotoxin B subunit from E. coli (LTB) and four epitopes from PRRSV. LTB-PRRSV was rescued as inclusion bodies and methods for its solubilization, IMAC-based purification, and refolding were standardized, leading to mean yields of 18 mg of pure protein per liter culture. Layered double hydroxides (LDH) have been used as vaccine adjuvants given their biocompatibility, low cost, and positive surface charge that allows an efficient adsorption of negatively charged biomolecules. Therefore, LDH were selected as delivery vehicles of LTB-PRRSV. Pure LTB-PRRSV was adsorbed onto LDH by incubation at different LDH:LTB-PRRSV mass ratios (1:0.25, 1:0.5, 1:1, and 1:2) and at pH 9.5. The best adsorption occurred with a 1:2 mass ratio, and in a sucrose-tween solution. The conjugates obtained had a polydispersity index of 0.26, a hydrodynamic diameter of 192 nm, and a final antigen concentration of 64.2 µg/mL. An immunogenicity assessment was performed by injecting mice with LDH:LTB-PRRSV, Alum/LTB-PRRSV, or LTB-PRRSV in a scheme comprising three immunizations at two-week intervals and two dose levels (1 and 5 µg). LTB-PRRSV was capable of inducing strong humoral responses, which lasted for a longer period when LDH was used as the delivery vehicle/adjuvant. The potential of LDH to serve as an attractive carrier for veterinary vaccines is discussed.