Kidney Immune Cell Characterization of Humanized Mouse Models.

Experimental studies often fail to translate to clinical practice. Humanized mouse models are an important tool to close this gap. We immunophenotyped the kidneys of NOG (EXL) and NSG mouse strains engrafted with human CD34 + hematopoietic stem cells or PBMCs and compared with immune cell composition of normal human kidney. Human CD34 + hematopoietic stem cell engraftment results in steady renal immune cell populations in mouse kidney with key similarities in composition compared with human kidney. Successful translation of experimental mouse data to human diseases is limited because of biological differences and imperfect disease models. Humanized mouse models are being used to bring murine models closer to humans. However, data for application in renal immune cell-mediated diseases are rare. We therefore studied immune cell composition of three different humanized mouse kidneys and compared them with human kidney. NOG and NOGEXL mice engrafted with human CD34 + hematopoietic stem cells were compared with NSG mice engrafted with human PBMCs. Engraftment was confirmed with flow cytometry, and immune cell composition in kidney, blood, spleen, and bone marrow was analyzed in different models. The results from immunophenotyping of kidneys from different humanized mouse strains were compared with normal portions of human kidneys. We found significant engraftment of human immune cells in blood and kidney of all tested models. huNSG mice showed highest frequencies of hTCR + cells compared with huNOG and huNOGEXL in blood. huNOGEXL was found to have the highest hCD4 + frequency among all tested models. Non-T cells such as hCD20 + and hCD11c + cells were decreased in huNSG mice compared with huNOG and huNOGEXL. Compared with normal human kidney, huNOG and huNOGEXL mice showed representative immune cell composition, rather than huNSG mice. In summary, humanization results in immune cell infiltration in the kidney with variable immune cell composition of tested humanized mouse models and partially reflects normal human kidneys, suggesting potential use for translational studies.

Renal tubular epithelial cells are constitutive non-cognate stimulators of resident T cells.

Understanding the roles of different cell types in regulating T cell homeostasis in various tissues is critical for understanding adaptive immunity. Here, we show that RTECs (renal tubular epithelial cells) are intrinsically programmed to polyclonally stimulate proliferation of kidney αß T cells by a cell-cell contact mechanism that is major histocompatibility complex (MHC) independent and regulated by CD155, αVß3-integrin, and vitronectin. Peripheral CD4 and CD8 are resistant to RTEC-mediated stimulation, while the minor subset of double-negative (DN) T cells are responsive. This functional property of RTEC is discovered by using a coculture system that recapitulates spontaneous in vivo polyclonal proliferation of kidney T cells, which are mainly comprised of central memory T (TCM) and effector memory T (TEM) cells. This robust cell-intrinsic stimulatory role of RTECs could be underlying the steady-state spontaneous proliferation of kidney T cells. The results have conceptual implications for understanding roles of different cell types in regulating systemic and organ-specific T cell homeostasis.

T Cell Nrf2/Keap1 Gene Editing Using CRISPR/Cas9 and Experimental Kidney Ischemia-Reperfusion Injury.

Aims: T cells play pathophysiologic roles in kidney ischemia-reperfusion injury (IRI), and the nuclear factor erythroid 2-related factor 2/kelch-like ECH-associated protein 1 (Nrf2/Keap1) pathway regulates T cell responses. We hypothesized that clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9)-mediated Keap1-knockout (KO) augments Nrf2 antioxidant potential of CD4+ T cells, and that Keap1-KO CD4+ T cell immunotherapy protects from kidney IRI. Results: CD4+ T cell Keap1-KO resulted in significant increase of Nrf2 target genes NAD(P)H quinone dehydrogenase 1, heme oxygenase 1, glutamate-cysteine ligase catalytic subunit, and glutamate-cysteine ligase modifier subunit. Keap1-KO cells displayed no signs of exhaustion, and had significantly lower levels of interleukin 2 (IL2) and IL6 in normoxic conditions, but increased interferon gamma in hypoxic conditions in vitro. In vivo, adoptive transfer of Keap1-KO CD4+ T cells before IRI improved kidney function in T cell-deficient nu/nu mice compared with mice receiving unedited control CD4+ T cells. Keap1-KO CD4+ T cells isolated from recipient kidneys 24 h post IR were less activated compared with unedited CD4+ T cells, isolated from control kidneys. Innovation: Editing Nrf2/Keap1 pathway in murine T cells using CRISPR/Cas9 is an innovative and promising immunotherapy approach for kidney IRI and possibly other solid organ IRI. Conclusion: CRISPR/Cas9-mediated Keap1-KO increased Nrf2-regulated antioxidant gene expression in murine CD4+ T cells, modified responses to in vitro hypoxia and in vivo kidney IRI. Gene editing targeting the Nrf2/Keap1 pathway in T cells is a promising approach for immune-mediated kidney diseases.

Sports injury and illness incidence among South Korean elite athletes in the 2018 Asian Games: a single-physician prospective study of 782 athletes.

OBJECTIVE: The increasing incidence of sports injury among athletes calls for systemic surveillance of injuries and illnesses in this field to develop preventive measures. The patterns of injuries and illnesses that occurred among Korean athletes during the 2018 Asian Games held in Indonesia were studied. METHODS: We recorded the occurrence of all injuries and illnesses reported to the chief medical officer, coordinated with the help of an instant social messaging application in real time. RESULTS: A total of 782 elite athletes participated in 46 sporting events. A total of 141 (18.03%) injuries were recorded, with 121 (15.47%) athletes suffering at least one injury. Out of 141 injuries 80 (56.74%) were in male athletes and 61 (43.26%) were in female athletes. The highest number of injuries was seen among sport climbing athletes (n=10, 71.43%), followed by sepak takraw. A total of 16 (11.35%) injuries were expected to prevent athletes from participation in competition/training. Most of the injuries occurred during training (46.10%), with lower lumbar spine being the most common part injured. A total of 209 (26.72%) illnesses were reported, with at least one illness in 170 (21.73%) athletes. The incidence among female athletes (26.90%) was comparable with that of male athletes (26.90%). Maximum illness rate was reported in table tennis (100%). The most common system involved was gastrointestinal (n=93, 44.49%), followed by respiratory (n=53, 25.36%). Environmental factors were causative in 111 athletes (53.11%) and infection in 79 (37.79%). Illnesses resulted in loss of at least 1 day among 30 (14.35%) athletes. CONCLUSION: Overall 15.47% of athletes suffered at least one injury and 21.73% suffered at least one illness; the incidence of injury and illness varied depending on the type of sports.

Persistent Interferon Production by Double Negative T Cells and Collapsing Focal Segmental Glomerulosclerosis.

Collapsing glomerulopathy has multiple associations, including viral infections, medications like bisphosphonates and interferon, autoimmune diseases, and genetic predisposition. We report a case of collapsing focal segmental glomerulosclerosis associated with persistently high levels of interferon gamma produced by T-cell receptor αß (+), CD4- CD8- (double negative) T lymphocytes that progressed despite treatment and improvement of other cytokine levels. Double negative T cells are elevated and activated in autoimmune lymphoproliferative syndrome (ALPS). Production of elevated interferon gamma levels from double negative T cells in ALPS despite treatment provides insight to the pathophysiology of collapsing glomerulopathy, guiding future research for collapsing glomerulopathy.

Lack of Syndecan-1 produces significant alterations in whole-body composition, metabolism and glucose homeostasis in mice.

BACKGROUND: Obesity is a disease state with serious adverse metabolic complications, including glucose intolerance and type 2 diabetes that currently has no cure. Identifying and understanding roles of various modulators of body composition and glucose homeostasis is required for developing effective cures. Syndecan-1 (Sdc1) is a member of the heparan sulfate proteoglycan family that has mainly been investigated for its role in regulating proliferation and survival of epithelia and tumor cells, but little is known about its roles in regulating obesity and glucose homeostasis. AIM: To examine the role of Sdc1 in regulating body fat and glucose metabolism. METHODS: We used female wild type and Sdc1 knockout (Sdc1 KO) mice on BALB/c background and multiple methods. Metabolic measurements (rates of oxygen consumption, carbon dioxide production, respiratory exchange ratio and energy expenditure) were performed using an open-flow indirect calorimeter with additional features to measure food intake and physical activity. Glucose intolerance and insulin resistance were measured by established tolerance test methods. RESULTS: Although our primary goal was to investigate the effects of Sdc1 deficiency on body fat and glucose homeostasis, we uncovered that Sdc1 regulates multiple metabolic parameters. Sdc1KO mice have reduced body weight due to significant decreases in fat and lean masses under both chow and high fat diet conditions. The reduced body weight was not due to changes in food intakes, but Sdc1 KO mice exhibited altered feeding behavior as they ate more during the dark phase and less during the light phase than wild type mice. In addition, Sdc1 KO mice suffered from high rate of energy expenditure, glucose intolerance and insulin resistance. CONCLUSION: These results reveal critical multisystem and opposing roles for Sdc1 in regulating normal energy balance and glucose homeostasis. The results will have important implications for targeting Sdc1 to modulate metabolic parameters. Finally, we offer a novel hypothesis that could reconcile the opposing roles associated with Sdc1 deficiency.

T Lymphocytes in Acute Kidney Injury and Repair.

Innate and adaptive immune systems participate in the pathogenesis of acute kidney injury (AKI). Considerable data from different research teams have shown the importance of T lymphocytes in the pathophysiology of AKI and, more recently, prevention and repair. T cells can generate or resolve inflammation by secreting specific cytokines and growth factors as well as interact with other immune and stromal cells to induce kidney injury or promote tissue repair. There also are emerging data on the role of T cells in the progression of AKI to chronic kidney disease and organ cross-talk in AKI. These data set the stage for immunomodulatory therapies for AKI. This review focuses on the major populations of T lymphocytes and their roles as mediators for AKI and repair.

TCR+CD4-CD8- (double negative) T cells protect from cisplatin-induced renal epithelial cell apoptosis and acute kidney injury.

Acute kidney injury (AKI) due to cisplatin is a significant problem that limits its use as an effective chemotherapeutic agent. T cell receptor+CD4-CD8- double negative (DN) T cells constitute the major T cell population in the human and mouse kidney, express programmed cell death protein (PD)-1, and protect from ischemic AKI. However, the pathophysiological roles of DN T cells in cisplatin-induced AKI is unknown. In this study, wild-type mice were treated with cisplatin (30 mg/kg) or vehicle, and the effects on kidney DN T cell numbers and function were measured. In vitro experiments evaluated effects of kidney DN T cells on cisplatin-induced apoptosis and PD ligand 1 (PD-L1) in renal epithelial cells. Adoptive transfer experiments assessed the therapeutic potential of DN T cells during cisplatin-induced AKI. Our results show that kidney DN T cell population increased at 24 h and declined by 72 h after cisplatin treatment. Cisplatin treatment increased kidney DN T cell proliferation, apoptosis, CD69, and IL-10 expression, whereas CD62L, CD44, IL-17A, interferon-γ, and TNF-α were downregulated. Cisplatin treatment decreased both PD-1 and natural killer 1.1 subsets of kidney DN T cells with a pronounced effect on the PD-1 subset. In vitro kidney DN T cell coculture decreased cisplatin-induced apoptosis in kidney proximal tubular epithelial cells, increased Bcl-2, and decreased cleaved caspase 3 expression. Cisplatin-induced expression of PD ligand 1 was reduced in proximal tubular epithelial cells cocultured with DN T cells. Adoptive transfer of DN T cells attenuated kidney dysfunction and structural damage from cisplatin-induced AKI. These results demonstrate that kidney DN T cells respond rapidly and play a protective role during cisplatin-induced AKI.

CD4+ T Cell-Derived NGAL Modifies the Outcome of Ischemic Acute Kidney Injury.

CD4+ T cells mediate the pathogenesis of ischemic and nephrotoxic acute kidney injury (AKI). However, the underlying mechanisms of CD4+ T cell-mediated pathogenesis are largely unknown. We therefore conducted unbiased RNA-sequencing to discover novel mechanistic pathways of kidney CD4+ T cells after ischemia compared with normal mouse kidney. Unexpectedly, the lipocalin-2 (Lcn2) gene, which encodes neutrophil gelatinase-associated lipocalin (NGAL) had the highest fold increase (∼60). The NGAL increase in CD4+ T cells during AKI was confirmed at the mRNA level with quantitative real-time PCR and at the protein level with ELISA. NGAL is a potential biomarker for the early detection of AKI and has multiple potential biological functions. However, the role of NGAL produced by CD4+ T cells is not known. We found that ischemic AKI in NGAL knockout (KO) mice had worse renal outcomes compared with wild-type (WT) mice. Adoptive transfer of NGAL-deficient CD4+ T cells from NGAL KO mice into CD4 KO or WT mice led to worse renal function than transfer of WT CD4+ T cells. In vitro-simulated ischemia/reperfusion showed that NGAL-deficient CD4+ T cells express higher levels of IFN-γ mRNA compared with WT CD4+ T cells. In vitro differentiation of naive CD4+ T cells to Th17, Th1, and Th2 cells led to significant increase in Lcn2 expression. Human kidney CD4+ T cell NGAL also increased significantly after ischemia. These results demonstrate an important role for CD4+ T cell NGAL as a mechanism by which CD4+ T cells mediate AKI and extend the importance of NGAL in AKI beyond diagnostics.

Gold nanocubes embedded biocompatible hybrid hydrogels for electrochemical detection of H2O2.

Smart electrochemical biosensors have emerged as a promising alternative analytical diagnostic tool in recent clinical practice. However, improvement in the biocompatibility and electrical conductivity of the biosensor matrix and the immobilization of various bioactive molecules such as enzymes still remain challenging. The present research reports the synthesis of a biocompatible hydrogel network and its integration with gold nanocubes (AuNCs) for developing a novel biosensor with improved functionality. The interpenetrating hydrogel network consist of biopolymers developed using graft co-polymerization of ß-cyclodextrin (ß-CD) and chitosan (CS). The novelty of this work is in integrating the CS-g-ß-CD hydrogel network with conductive AuNCs for improving hydrogel conductivity, biosensor sensitivity and use of the material for a biocompatible sensor. The present protocol advances the state of the art for the utilization of biopolymeric hydrogels system in synergy with an enzymatic biosensing protocol for exclusively detecting hydrogen peroxide (H2O2). Immobilization of the mitochondrial protein, cytochrome c (cyt c) into the hydrogel nanocomposite matrix was performed via thiol cross-linking. This organic-inorganic hybrid nanocomposite hydrogel matrix exhibited high biocompatibility (RAW 264.7 and N2a cell lines), improved electrical conductivity to attain high sensitivity (1.2 mA mM-1 cm-2) and a low detection limit (15 × 10-9 M) for H2O2.

Hybrid lipids, peptides, and lymphocytes: new era in type 1 diabetes research.

Type 1 diabetes (T1D) results from autoimmune destruction of insulin-producing ß cells in islets of Langerhans. Many genetic and immunological insights into autoimmune disease pathogenesis were initially uncovered in the context of T1D and facilitated by preclinical studies using the nonobese diabetic (NOD) mouse model. Recently, the study of T1D has led to the discovery of fatty acid esters of hydroxyl fatty acids (FAHFAs), which are naturally occurring hybrid peptides that modulate inflammation and diabetes pathogenesis, and a hybrid lymphocyte that expresses both B and T cell receptors. Palmitic acid esters of hydroxy stearic acids (PAHSAs) are the most extensively studied FAHFA. In this issue of the JCI, Syed et al. have shown that PAHSAs both attenuate autoimmune responses and promote ß cell survival in NOD mice. Given the lack of effective T1D therapies and the paucity of known side effects of PAHSAs, this lipid may have therapeutic potential for individuals at risk for or newly diagnosed with T1D.

Activation and Proliferation of PD-1+ Kidney Double-Negative T Cells Is Dependent on Nonclassical MHC Proteins and IL-2.

BACKGROUND: CD4- CD8- double-negative (DN) αß T cells with innate-like properties represent a significant component of T cells in human and mouse kidneys. They spontaneously proliferate in the steady state and protect against ischemic AKI. However, the mechanisms regulating DN T cell homeostasis and responses to external danger signals from "sterile" inflammation remain poorly understood. METHODS: We used knockout mice, functional assays, and an established ischemic AKI model to investigate the role of various MHC class I and II molecules in regulating kidney DN T cells. We also studied human nephrectomy samples. RESULTS: Deficiency of ß2m-dependent MHC class I (but not MHC class II) molecules led to significant reduction in frequency or absolute numbers of kidney DN T cells due to impaired activation, proliferation, increased apoptosis, and loss of an NK1.1+ subset of DN T cells. The remaining DN T cells in ß2m knockout mice mainly comprised a programmed cell death protein-1 receptor (PD-1+) subset that depends on IL-2 provided by conventional T cells for optimal homeostasis. However, this PD-1+ subset remained highly responsive to changes in milieu, demonstrated by responses to infused lymphocytes. It was also the major responder to ischemic AKI; the NK1.1+ subset and CD8+ T cells had minimal responses. We found both DN T cell subsets in normal and cancerous human kidneys, indicating possible clinical relevance. CONCLUSIONS: DN T cells, a unique population of kidney T cells, depend on nonclassical ß2m molecules for homeostasis and use MHC-independent mechanisms to respond to external stimuli. These results have important implications for understanding the role these cells play during AKI and other immune cell-mediated kidney diseases.

Unexpected alliance between syndecan-1 and innate-like T cells to protect host from autoimmune effects of interleukin-17.

Innate-like T cells, namely natural killer T (NKT) and γδ T cells, play critical roles in linking innate and adaptive immune responses through rapid production of cytokines. Prominent among these cytokines is interleukin-17 (IL-17), which is a potent proinflammatory cytokine that plays a critical role in host defense against fungi and extracellular bacteria. However, excessive IL-17-production promotes autoimmune diseases, including psoriasis, multiple sclerosis, rheumatoid arthritis, inflammatory bowel disease, and systemic lupus erythematosus. IL-17 has also been implicated in regulating body fat, which is highly relevant given rises in obesity and type 2 diabetes. NKT cells, γδ T cells and mucosal-associated invariant T cells (MAIT) are the major sources of IL-17 involved in protection of mucosal surfaces from opportunistic infections and causing autoimmunity when become dysregulated. Given the pathogenic effects of IL-17, efforts have been directed towards understanding mechanisms that guard against IL-17 overproduction. One novel potent mechanism is mediated by the heparan sulfate proteoglycan, syndecan-1 (sdc1), which is selectively expressed by IL-17-producing subsets of NKT and γδ T cells. This unexpected role for sdc1 is uncovered by analysis of NKT and γδ T cells in sdc1-deficient mice. In this mini-review, we discuss selective expression of sdc1 by these innate T cells and consequences of its absence on IL-17 homeostasis and pathological implications.

Syndecan-1 Regulates Psoriasiform Dermatitis by Controlling Homeostasis of IL-17-Producing γδ T Cells.

IL-17 is a potent proinflammatory cytokine that drives pathogenesis of multiple autoimmune diseases, including psoriasis. A major source of pathogenic IL-17 is a subset of γδ T cells (Tγδ17) that acquires the ability to produce IL-17 while developing in the thymus. The mechanisms that regulate homeostasis of Tγδ17 cells and their roles in psoriasis, however, are not fully understood. In this paper, we show that the heparan sulfate proteoglycan syndecan-1 (sdc1) plays a critical role in regulating homeostasis of Tγδ17 cells and modulating psoriasis-like skin inflammation in mice. sdc1 was predominantly expressed by Tγδ17 cells (but not IL-17- Tγδ cells) in the thymus, lymph nodes, and dermis. sdc1 deficiency significantly and selectively increased the frequency and absolute numbers of Tγδ17 cells by mechanisms that included increased proliferation and decreased apoptosis. Adoptive transfer experiments ruled out a significant role of sdc1 expressed on nonhematopoietic cells in halting expansion and proliferation of sdc1-deficient Tγδ17 cells. When subjected to imiquimod-induced psoriasiform dermatitis, Tγδ17 cells in sdc1KO mice displayed heightened responses accompanied by significantly increased skin inflammation than their wild-type counterparts. Furthermore, transferred sdc1-deficient γδ T cells caused more severe psoriasiform dermatitis than their sdc1-sufficient counterparts in TCR-ßδ KO hosts. The results uncover a novel role for sdc1 in controlling homeostasis of Tγδ17 cells and moderating host responses to psoriasis-like inflammation.

Characterization of kidney CD45intCD11bintF4/80+MHCII+CX3CR1+Ly6C- "intermediate mononuclear phagocytic cells".

Kidney immune cells play important roles in pathogenesis of many diseases, including ischemia-reperfusion injury (IRI) and transplant rejection. While studying murine kidney T cells, we serendipitously identified a kidney mononuclear phagocytic cell (MPC) subset characterized by intermediate surface expression of CD45 and CD11b. These CD45intCD11bint MPCs were further identified as F4/80+MHCII+CX3CR1+Ly6C- cells, comprising ~17% of total CD45+ cells in normal mouse kidney (P < 0.01) and virtually absent from all other organs examined except the heart. Systemic clodronate treatment had more significant depletive effect on the CD45intCD11bint population (77.3%±5.9%, P = 0.03) than on CD45highCD11b+ population (14.8%±16.6%, P = 0.49). In addition, CD45intCD11bint MPCs had higher phagocytic function in the normal kidney (35.6%±3.3% vs. 24.1%±2.2%, P = 0.04), but lower phagocytic capacity in post-ischemic kidney (54.9%±1.0% vs. 67.8%±1.9%, P < 0.01) compared to the CD45highCD11b+ population. Moreover, the CD45intCD11bint population had higher intracellular production of the pro-inflammatory tumor necrosis factor (TNF)-α (58.4%±5.2% vs. 27.3%±0.9%, P < 0.001) after lipopolysaccharide (LPS) stimulation and lower production of the anti-inflammatory interleukin (IL)-10 (7.2%±1.3% vs. 14.9%±2.2%, P = 0.02) following kidney IRI, suggesting a functional role under inflammatory conditions. The CD45intCD11bint cells increased early after IRI, and then abruptly decreased 48h later, whereas CD45highCD11b+ cells steadily increased after IRI before declining at 72h (P = 0.03). We also identified the CD45intCD11bint MPC subtype in human kidney. We conclude that CD45intCD11bint F4/80+MHCII+CX3CR1+Ly6C-population represent a unique subset of MPCs found in both mouse and human kidneys. Future studies will further characterize their role in kidney health and disease.

KEAP1 Editing Using CRISPR/Cas9 for Therapeutic NRF2 Activation in Primary Human T Lymphocytes.

Oxidant stress modifies T lymphocyte activation and function. Previous work demonstrated that murine T cell-specific kelch like-ECH-associated protein 1 (Keap1) deletion enhances antioxidant capacity and protects from experimental acute kidney injury. In this study, we used CRISPR technology to develop clinically translatable human T cell-specific KEAP1 deletion. Delivery of KEAP1 exon 2 specific Cas9:guide RNA in Jurkat T cells led to significant (∼70%) editing and upregulation of NRF2-regulated antioxidant genes NADPH dehydrogenase quinone 1 (NQO1) (up to 11-fold), heme oxygenase 1 (HO1) (up to 11-fold), and GCLM (up to 2-fold). In primary human T cells, delivery of KEAP1 exon 2 target site 2-specific ATTO 550-labeled Cas9:guide RNA edited KEAP1 in ∼40% cells and significantly (p ≤ 0.04) increased NQO1 (16-fold), HO1 (9-fold), and GCLM (2-fold) expression. To further enrich KEAP1-edited cells, ATTO 550-positive cells were sorted 24 h after electroporation. Assessment of ATTO 550-positive cells showed KEAP1 editing in ∼55% cells. There was no detectable off-target cleavage in the top three predicted genes in the ATTO 550-positive cells. Gene expression analysis found significantly (p ≤ 0.01) higher expression of NQO1 mRNA in ATTO 550-positive cells compared with control cells. Flow cytometric assessment showed increased (p ≤ 0.01) frequency of CD4-, CD25-, and CD69-expressing KEAP1 edited cells whereas frequency of CD8- (p ≤ 0.01) and IL-17- (p ≤ 0.05) expressing cells was reduced compared with control cells. Similar experimental conditions resulted in significant KEAP1 editing, increased antioxidant gene expression, and frequency of CD69 and IL-10 positive cells in highly enriched KEAP1-edited regulatory T cells. KEAP1-edited T cells could potentially be used for treating multiple human diseases.

Role of Immune Cells in Acute Kidney Injury and Repair.

Acute kidney injury (AKI) is a significant problem in both native and transplant kidneys. There have been significant advances in understanding the role of immune cells in the early injury and repair from AKI. In this brief review, we aim to update information on the pathophysiologic impact of various immune cells in AKI, with special emphasis on repair. An improved understanding of the AKI immunopathology will lead to new therapies that prevent AKI, accelerate repair, and prevent the progression of AKI to chronic kidney disease.

Syndecan-1-coating of interleukin-17-producing natural killer T cells provides a specific method for their visualization and analysis.

Natural killer T cells (NKT cells) are innate-like T cells that acquire effector functions while developing in the thymus, polarize into three distinct functional subsets viz. NKT1, NKT2 and NKT17 cells that produce interferon (IFN)-γ, interleukin (IL)-4 and IL-17, respectively. However, there has been no unique surface markers that define each subsets, forcing investigators to use intracellular staining of transcription factors and cytokines in combination of surface markers to distinguish among these subsets. Intracellular staining, however, causes apoptosis and prevents subsequent utilization of NKT cells in functional in vitro and in vivo assays that require viable cells. This limitation has significantly impeded understanding the specific properties of each subset and their interactions with each other. Therefore, there has been fervent efforts to find a specific markers for each NKT cell subset. We have recently identified that syndecan-1 (SDC-1; CD138) as a specific surface marker of NKT17 cells. This discovery now allows visualization of NKT17 in situ and study of their peripheral tissue distribution, characteristics of their TCR and viable sorting for in vitro and in vivo analysis. In addition, it lays the ground working for investigating significance of SDC-1 expression on this particular subset in regulating their roles in host defense and glucose metabolism.

Sirtuin 4 Regulates Lipopolysaccharide Mediated Leydig Cell Dysfunction.

Bacterial lipopolysaccharide (LPS) is the most important contributing factor in pathogenesis of bacterial infection in male accessory glands; and it has shown to inhibit testicular steroidogenesis and induce apoptosis. The present study demonstrates that LPS causes mitochondrial dysfunction via suppression of sirtuin 4 (SIRT4); which in turn affects Leydig cell function by modulating steroidogenesis and apoptosis. LC-540 Leydig cells treated with LPS (10 µg/ml) showed impaired steroidogenesis and increased cellular apoptosis. The mRNA and protein expression of SIRT4 were decreased in LPS treated cells when compared to controls. The obtained data suggest that the c-Jun N-terminal kinase (JNK) activation suppresses SIRT4 expression in LPS treated Leydig cells. Furthermore, the overexpression of SIRT4 prevented LPS induced impaired steroidogenesis and cellular apoptosis by improving mitochondrial function. These findings provide valuable information that SIRT4 regulates LPS mediated Leydig cell dysfunction.