Transient receptor potential channel 6 (TRPC6) protects podocytes during complement-mediated glomerular disease.

Gain-of-function mutations in the calcium channel TRPC6 lead to autosomal dominant focal segmental glomerulosclerosis and podocyte expression of TRPC6 is increased in some acquired human glomerular diseases, particularly in membranous nephropathy. These observations led to the hypothesis that TRPC6 overactivation is deleterious to podocytes through pathological calcium signaling, both in genetic and acquired diseases. Here, we show that the effects of TRPC6 on podocyte function are context-dependent. Overexpression of TRPC6 alone did not directly affect podocyte morphology and cytoskeletal structure. Unexpectedly, however, overexpression of TRPC6 protected podocytes from complement-mediated injury, whereas genetic or pharmacological TRPC6 inactivation increased podocyte susceptibility to complement. Mechanistically, this effect was mediated by Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) activation. Podocyte-specific TRPC6 transgenic mice showed stronger CaMKII activation, reduced podocyte foot process effacement and reduced levels of proteinuria during nephrotoxic serum nephritis, whereas TRPC6 null mice exhibited reduced CaMKII activation and higher levels of proteinuria compared with wild type littermates. Human membranous nephropathy biopsy samples showed podocyte staining for active CaMKII, which correlated with the degree of TRPC6 expression. Together, these data suggest a dual and context dependent role of TRPC6 in podocytes where acute activation protects from complement-mediated damage, but chronic overactivation leads to focal segmental glomerulosclerosis.

Prevalence of Sensory Dysfunction in Smell and/or Taste in Veterans with Traumatic Brain Injury and Link to Demographics and Comorbidities.

INTRODUCTION: Traumatic brain injury (TBI) entails chronic neurological symptoms and deficits, such as smell and taste impairment. In the general population, a prevalence of 13.5% for smell impairment, 17% for taste impairment, and 2.2% for both have been reported. Studies establishing prevalence of sensorial dysfunction in the U.S. general population showed that prevalence increased with age and was higher in ethnic minorities and men. To understand the mechanisms that lead to these deficits, the prevalence of sensory dysfunction was studied in the Veteran TBI population of the VA Caribbean Healthcare System (VACHS). The aims were to find the prevalence of sensorial dysfunction in smell and/or taste in TBI patients at the VACHS Polytrauma Clinic and its association with demographic characteristics and medical comorbidities. The hypothesis was that the prevalence of sensory dysfunction in smell and/or taste of VACHS Veterans (mostly Hispanics minority) with TBI will be higher than the one historically reported in the literature for the U.S. general population. MATERIALS AND METHODS: A retrospective record review was held at the VACHS Polytrauma Clinic from January 2018 to January 2020 (before coronavirus disease 2019 pandemic) to evaluate the prevalence of sensory dysfunction. Data on demographics and comorbidities in the electronic medical records, and the TBI Second-Level Evaluation note, that was previously completed by a physician from the Polytrauma Clinic at the VACHS to diagnose and characterized the TBI event, were reviewed. Data were summarized using descriptive statistics. To establish the relation among demographic characteristics and comorbidities with the prevalence of smell and/or taste sensory dysfunction, chi-square and Fisher's exact tests were used. RESULTS: A total of 81 records were reviewed. This corresponded to all the patients diagnosed with TBI in the VACHS Polytrauma Clinic from January 2018 to January 2020. The prevalence of sensory dysfunction in the studied population was 38.3%. Men tend to present a higher prevalence of smell and/or taste dysfunction (40.0%) in comparison with women (16.7%); however, the difference did not achieve statistical significance (P = .399). Hispanics had a relatively higher prevalence of sensory dysfunction than non-Hispanics, but this difference did not reach statistical significance (P = .210). Forty-nine subjects were combat Veterans (60.5%). There was a significant correlation regarding the combat status of the subjects (P = .014), where 24 of the 49 combat Veterans presented smell and/or taste dysfunction (49.0%). A marginal significance was observed for obesity; obese participants were less likely to have a significant smell and/or taste dysfunction (P = .053). CONCLUSION: The investigators found that the prevalence of sensory dysfunction in smell and/or taste in VACHS Veterans with TBI was 38.3% (n = 31). A significant association was found between smell and/or taste dysfunction and being a combat veteran (P = .018). A marginally significant association to obesity was also observed (P = .053). To the scientific community, the results will serve as a base for sensorial dysfunction and TBI research given that this prevalence, and the correlation to demographics and comorbidities, has not been fully established in the Veteran population.

Macrophage response to staphylococcal biofilms on crosslinked poly(ethylene) glycol polymer coatings and common biomaterials in vitro.

Biomaterial-associated-infections (BAI) are serious clinical complications that threaten the longevity of implanted devices and lead to high morbidity and mortality. Poly(ethylene)glycol (PEG) coatings have been studied as a strategy to reduce the incidence of BAI by reducing protein deposition that promotes pathogen adhesion and growth on device surfaces. Despite their effectiveness to reduce protein adsorption and a hundred-fold reduction in bacterial adhesion, PEG-based coatings still facilitate weak bacterial adhesion that can form an initial basis for biofilms. Here, we describe a methodology enabling direct, quantitative and detailed qualitative in situ observation of macrophage morphology, migration and phagocytosis of bacteria. In vitro interaction of macrophages with Staphylococcus epidermidis 3399 adhering to commercial, crosslinked PEG-based coatings (OptiChem®) was compared with fluorinated ethylene propylene, silicone rubber and glass. Adhesion, phagocytosis and migration were studied real-time in a parallel-plate-flow-chamber. Macrophages cultured on OptiChem® coatings showed enhanced migration and phagocytosis of bacteria compared to common biomaterials. Bacterial clearance per macrophage on both inert and reactive OptiChem® coatings were about three times higher than on the common biomaterials studied, corresponding with up to 70% reduction in bacterial numbers on OptiChem®, whereas on the biomaterials less than 40% bacterial reduction was obtained. These findings show that bacterial clearance from cross-linked PEG-based coatings by macrophages is more effective than from common biomaterials, possibly resulting from weak adhesion of bacteria on Optichem®. Moreover, macrophages exhibit higher mobility on Optichem® retaining an improved capability to clear bacteria from larger areas than from other common biomaterials, where they appear more immobilized.

Enhancing Language Access: A Pilot Study to Examine the Importance of Understanding the Language Preference and Acculturalization Level in the Provision of Healthcare for Hispanics Veterans With Traumatic Brain Injury.

INTRODUCTION: The purpose of this pilot study was to obtain preliminary data to culturally adapt the Veteran Health Administration Traumatic Brain Injury (TBI) assessment instruments for the Hispanic Veteran population. A qualitative analysis explored the cognitive processes used by Hispanic Veterans whose preferred language was Spanish to understand a specific set of screening questions within the Initial TBI Screening, the Comprehensive TBI Evaluation, the Neurobehavioral Symptom Inventory (NSI), and the La Trobe Communication Questionnaire (LTCQ). MATERIALS AND METHODS: A certified translator completed translation of the TBI instruments, an expert panel resolved inadequate expressions of the translations, and translated instruments were back translated. Male and female Hispanic Veterans with a positive TBI screening underwent a recorded administration of the TBI instruments, including LTCQ, followed by systematic debriefing using semi-structured cognitive interviews which then underwent qualitative analysis. The Marin's Short Acculturation Scale for Hispanics, the Tropp's Psychological Acculturation Scale, the English-Language Proficiency Test Series, and the TBI Demographic and Language Preference interview were administered to the subjects. RESULTS: Fifteen subjects were enrolled for the TBI instruments intervention; 11 of them completed all the additional procedures. The TBI instruments intervention seemed to produce very few variations, indicating adequate cultural equivalence. However, the LTCQ instrument showed suggested cultural variations, but did not suggest a lack of understanding or misinterpretation. The population studied displayed preferential connectedness to the Hispanic/Latino culture and to the Spanish language. The LTCQ indicated that subjects perceived themselves as having a worse execution in terms of communication skills than historical control and TBI groups. English-Language Proficiency Test Series found that most of the subject population did not demonstrate mastery of grade-appropriate basic social and academic vocabulary in English. CONCLUSION: Current findings highlight the importance of using linguistically and culturally appropriate materials upon evaluating Hispanic Veterans with a suspected TBI who have Spanish as their primary or preferred language.

The risk of biomaterial-associated infection after revision surgery due to an experimental primary implant infection.

The fate of secondary biomaterial implants was determined by bio-optical imaging and plate counting, after antibiotic treatment of biomaterials-associated-infection (BAI) and surgical removal of an experimentally infected, primary implant. All primary implants and tissue samples from control mice showed bioluminescence and were culture-positive. In an antibiotic treated group, no bioluminescence was detected and only 20% of all primary implants and no tissue samples were culture-positive. After revision surgery, bioluminescence was detected in all control mice. All the implants and 80% of all tissue samples were culture-positive. In contrast, in the antibiotic treated group, 17% of all secondary implants and 33% of all tissue samples were culture-positive, despite antibiotic treatment. The study illustrates that due to the BAI of a primary implant, the infection risk of biomaterial implants is higher in revision surgery than in primary surgery, emphasizing the need for full clearance of the infection, as well as from surrounding tissues prior to implantation of a secondary implant.

Bacterial colonization of polymer brush-coated and pristine silicone rubber implanted in infected pockets in mice.

OBJECTIVES: Curing biomaterial-associated infection (BAI) frequently includes antibiotic treatment, implant removal and re-implantation. However, revision implants are at a greater risk of infection as they may attract bacteria from their infected surroundings. Polymer brush-coatings attract low numbers of bacteria, but the virtue of polymer brush-coatings in vivo has seldom been investigated. Here, we determine the possible benefits of polymer brush-coated versus pristine silicone rubber in revision surgery, using a murine model. METHODS: BAI was induced in 26 mice by subcutaneous implantation of silicone rubber discs with a biofilm of Staphylococcus aureus Xen29. During the development of BAI, half of the mice received rifampicin/vancomycin treatment. After 5 days, the infected discs were removed from all mice, and either a polymer brush-coated or pristine silicone rubber disc was re-implanted. Revision discs were explanted after 5 days, and the number of cfu cultured from the discs and the surrounding tissue was determined. RESULTS: None of the polymer brush-coated discs after antibiotic treatment appeared colonized by staphylococci, whereas 83% of the pristine silicone rubber discs were re-infected. Polymer brush-coated discs also showed reduced colonization rates in the absence of antibiotic treatment when compared with pristine silicone rubber discs. Tissue surrounding the discs was culture-positive in all cases. CONCLUSIONS: Polymer brush-coatings are less prone to re-infection than pristine silicone rubber when used in revision surgery, i.e. when implanted in a subcutaneous pocket infected by a staphylococcal BAI. Antibiotic pre-treatment during the development of BAI hardly had any effect in preventing the colonization of pristine silicone rubber.

The inhibition of the adhesion of clinically isolated bacterial strains on multi-component cross-linked poly(ethylene glycol)-based polymer coatings.

This study examined bacterial adhesion to a new multi-component cross-linked poly(ethylene glycol)-based polymer coating that can be applied by spin-coating or spraying onto diverse biomaterials. Adhesion of five clinically isolated bacterial strains involved in biomaterial-centered infections were studied in a parallel-plate flow chamber at different shear rates and after exposure of the coating to different physiological fluids. The new chemistry inhibits non-specific biomolecular and cell binding interactions. Relative to glass, the coating reduced adhesion of all strains used in this study by more than 80%, with the exception of Escherichia coli O2K2. Reductions in adhesion of Staphylococcus epidermidis 3,399 persisted beyond 168h exposure of the coatings to phosphate buffered saline or urine, but not after exposure to protein-rich fluids as saliva and blood plasma, despite evidence from X-ray photoelectron spectroscopy that the coating integrity was not affected by exposure to these fluids. We conclude that this new coating chemistry provides beneficial properties to prevent or hinder bacterial adhesion and colonization in applications where low protein-conditions prevail.

Competitive time- and density-dependent adhesion of staphylococci and osteoblasts on crosslinked poly(ethylene glycol)-based polymer coatings in co-culture flow chambers.

Biomaterial-associated infections (BAI) remain a serious clinical complication, often arising from an inability of host tissue-implant integration to out-compete bacterial adhesion and growth. A commercial polymer coating based on polyethylene glycol (PEG), available in both chemically inert and NHS-activated forms (OptiChem(®)), was compared for simultaneous growth of staphylococci and osteoblasts. In the absence of staphylococci, osteoblasts adhered and proliferated well on glass controls and on the NHS-reactive PEG-based coating over 48 h, but not on the inert PEG coating. Staphylococcal growth was low on both PEG-based coatings. When staphylococci were pre-adhered on surfaces for 1.5 h to mimic peri-operative contamination, osteoblast growth and spreading was reduced on glass but virtually absent on both reactive and inert PEG-based coatings. Thus although NHS-reactive, PEG-based coatings stimulated tissue-cell interactions in the absence of contaminating staphylococci, the presence of adhering staphylococci eliminated osteoblast adhesion advantages on the PEG surface. This study demonstrates the importance of using bacterial and cellular co-cultures compared to monocultures when assessing functionalized biomaterials coatings for infectious potential.

In vitro interactions between bacteria, osteoblast-like cells and macrophages in the pathogenesis of biomaterial-associated infections.

Biomaterial-associated infections constitute a major clinical problem that is difficult to treat and often necessitates implant replacement. Pathogens can be introduced on an implant surface during surgery and compete with host cells attempting to integrate the implant. The fate of a biomaterial implant depends on the outcome of this race for the surface. Here we studied the competition between different bacterial strains and human U2OS osteoblast-like cells (ATCC HTB-94) for a poly(methylmethacrylate) surface in the absence or presence of macrophages in vitro using a peri-operative contamination model. Bacteria were seeded on the surface at a shear rate of 11 1/s prior to adhesion of U2OS cells and macrophages. Next, bacteria, U2OS cells and macrophages were allowed to grow simultaneously under low shear conditions (0.14 1/s). The outcome of the competition between bacteria and U2OS cells for the surface critically depended on bacterial virulence. In absence of macrophages, highly virulent Staphylococcus aureus or Pseudomonas aeruginosa stimulated U2OS cell death within 18 h of simultaneous growth on a surface. Moreover, these strains also caused cell death despite phagocytosis of adhering bacteria in presence of murine macrophages. Thus U2OS cells are bound to loose the race for a biomaterial surface against S. aureus or P. aeruginosa, even in presence of macrophages. In contrast, low-virulent Staphylococcus epidermidis did not cause U2OS cell death even after 48 h, regardless of the absence or presence of macrophages. Clinically, S. aureus and P. aeruginosa are known to yield acute and severe biomaterial-associated infections in contrast to S. epidermidis, mostly known to cause more low-grade infection. Thus it can be concluded that the model described possesses features concurring with clinical observations and therewith has potential for further studies on the simultaneous competition for an implant surface between tissue cells and pathogenic bacteria in presence of immune system components.

In vitro and in vivo comparisons of staphylococcal biofilm formation on a cross-linked poly(ethylene glycol)-based polymer coating.

Poly(ethylene glycol) (PEG) coatings are known to reduce microbial adhesion in terms of numbers and binding strength. However, bacterial adhesion remains of the order of 10(4)cm(-2). It is unknown whether this density of bacteria will eventually grow into a biofilm. This study investigates the kinetics of staphylococcal biofilm formation on a commercially produced, robust, cross-linked PEG-based polymer coating (OptiChem) in vitro and in vivo. OptiChem inhibits biofilm formation in vitro, and although adsorption of plasma proteins encourages biofilm formation, microbial growth kinetics are still strongly delayed compared to uncoated glass. In vivo, OptiChem-coated and bare silicone rubber samples were inserted into an infected murine subcutaneous pocket model. In contrast to bare silicone rubber, OptiChem samples did not become colonized upon reimplantation despite the fact that surrounding tissues were always culture-positive. We conclude that the commercial OptiChem coating considerably slows down bacterial biofilm formation both in vitro and in vivo, making it an attractive candidate for biomaterials implant coating.