Increased lacrimal inflammatory mediators in patients with keratoconus.

Purpose: This study aimed to characterize the tear film immunologic profile in keratoconus (KC) patients compared with healthy individuals (control group) and to investigate the correlation between the tear film immunologic profile and atopy, disease severity, and disease status over time. Methods: The study involved 30 KC patients and 18 healthy individuals. Tear collection was obtained using microcapillary tubes. Tear film levels of fractalkine, granulocyte-macrophage colony-stimulating factor (GM-CSF), interferon (IFN)-γ, interleukin (IL)-1ß, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12, IL-13, IL-17A, IL-21, IL-23, interferon-inducible T-cell alpha chemoattractant (ITAC), macrophage inflammatory protein-1 alpha (MIP-1α), MIP-1ß, MIP-3α, and tumor necrosis factor (TNF)-α were detected. Keratometric measurements and topographic patterns were used to diagnose and define disease progression. Tear immunologic profiles were compared, emphasizing the presence or absence of ocular allergy. Correlations between the cytokine profile, disease severity, and disease status were also analyzed longitudinally in the KC patients. Results: Lacrimal cytokine concentrations were higher in the KC patients than they were in the controls in 14 of 21 cytokines analyzed. IL-6 was the most relevant cytokine found in KC patients, especially when associated with ocular allergy. There was no correlation between KC progression and the level of inflammatory cytokines when analyzed longitudinally. KC severity correlated with IL-6 concentration, where the more severe KC presented a higher IL-6 concentration in tears. Conclusions: Inflammatory activity seems to be involved in the pathogenesis of KC. Out of 21 cytokines, 14 were more concentrated in the tears of KC patients than healthy subjects. IL-6 was significantly higher in KC patients' tears and was related to disease severity. Disease progression did not correlate with cytokine levels when analyzed longitudinally.

NF-κB System Is Chronically Activated and Promotes Glomerular Injury in Experimental Type 1 Diabetic Kidney Disease.

High glucose concentration can activate TLR4 and NF-κB, triggering the production of proinflammatory mediators. We investigated whether the NF-κB pathway is involved in the pathogenesis and progression of experimental diabetic kidney disease (DKD) in a model of long-term type 1 diabetes mellitus (DM). Adult male Munich-Wistar rats underwent DM by a single streptozotocin injection, and were kept moderately hyperglycemic by daily insulin injections. After 12 months, two subgroups - progressors and non-progressors - could be formed based on the degree of glomerulosclerosis. Only progressors exhibited renal TLR4, NF-κB and IL-6 activation. This scenario was already present in rats with short-term DM (2 months), at a time when no overt glomerulosclerosis can be detected. Chronic treatment with the NF-κB inhibitor, pyrrolidine dithiocarbamate (PDTC), prevented activation of renal TLR4, NF-κB or IL-6, without interfering with blood glucose. PDTC prevented the development of glomerular injury/inflammation and oxidative stress in DM rats. In addition, the NF-κB p65 component was detected in sclerotic glomeruli and inflamed interstitial areas in biopsy material from patients with type 1 DM. These observations indicate that the renal NF-κB pathway plays a key role in the development and progression of experimental DKD, and can become an important therapeutic target in the quest to prevent the progression of human DKD.

Gut Microbiota and Intestinal Epithelial Myd88 Signaling Are Crucial for Renal Injury in UUO Mice.

Increasing evidence shows the essential participation of gut microbiota in human health and diseases by shaping local and systemic immunity. Despite an accumulating body of studies showing that chronic kidney disease (CKD) is closely associated with disturbances in the composition of gut microbiota, it remains unclear the importance of gut microbiota in the onset and development of CKD. For the purpose of untangling the role of gut microbiota in CKD, gut microbiota was depleted with a pool of broad-spectrum antibiotics in mice submitted to unilateral ureteral obstruction (UUO). Depletion of gut microbiota significantly decreased levels of proinflammatory cytokines and fibrosis markers, attenuating renal injury. Additionally, to study whether the pathogenic role of gut microbiota is dependent of microbial-host crosstalk, we generated mice lacking Myd88 (myeloid differentiation primary response gene 8) expression in intestinal epithelial cells (IECs) and performed UUO. The absence of Myd88 in IECs prevented a bacterial burden in mesenteric lymph nodes as observed in WT mice after UUO and led to lower expression of proinflammatory cytokines and chemokines, reducing deposition of type I collagen and, ultimately, attenuating renal damage. Therefore, our results suggest that the presence of gut microbiota is crucial for the development of CKD and may be dependent of Myd88 signaling in IECs, which appears to be essential to maturation of immune cells intimately involved in aggravation of inflammatory scenarios.

l-Arginine supplementation blunts resistance exercise improvement in rats with chronic kidney disease.

Chronic kidney disease (CKD) patients present L-arginine (L-arg) deficiency and L-arg supplementation has been used as a treatment. In addition, sarcopenia is another common problem in CKD population, resistance training (RT) is one of the conservative strategies developed to prevent CKD progression, and however there are no evidences of a combination of these two strategies to treat CKD outcomes. The aim of this study was to evaluate the effects of oral L-arg supplementation combined with RT in an experimental model of CKD. Twenty-five Munich-Wistar male rats, 8-week-old were divided in 5 groups: Sham (sedentary control), Nx (CKD sedentary), Nx L-arg (CKD sedentary supplemented with 2% of L-arg), Nx RT (CKD exercised) Nx RT + L-arg (CKD exercised and supplemented with 2% of L-arg). CKD model was obtained by a subtotal 5/6 nephrectomy. RT was performed on a ladder climbing, three weekly sessions on non-consecutive days, with an intensity of 70% maximum carrying capacity. They were submitted to RT and/or L-arg supplementation for 10 weeks. There was a significant improvement in muscle strength, renal function, anti-inflammatory cytokines, arginase metabolism and renal fibrosis after RT. However, the combination of RT and L-arg impaired all the improvements promoted by RT alone. The L-arg supplementation alone did not impair renal fibrosis and renal function. In conclusion, RT improved inflammatory balance, muscle strength, renal function and consequently decreased renal fibrosis. Nevertheless, the association with L-arg supplementation prevented all these effects promoted by RT.

Resistance training downregulates macrophages infiltration in the kidney of 5/6 nephrectomized rats.

INTRODUCTION: Chronic kidney disease (CKD) is considered a significant world health problem with elevated mortality rates. Patients with CKD are restricted to mild physical activity, present chronic inflammatory state and loss of muscle strength. Currently, the influence of resistance exercise (RE) on the progression of renal disease has not being fully elucidated. PURPOSE: To evaluate the effects of RE on the progression of CKD in a remnant kidney model (5/6Nx) in rats. METHODS: Eight-week-old Wistar rats were submitted to 5/6 nephrectomy and were divided into four groups: Sham sedentary (Sham SD); Sham RE (Sham RE); 5/6Nx SD and 5/6Nx RE. The animals were trained for 8 weeks in a vertical climbing ladder for 3 days per week, on non-consecutive days. RESULTS: As expected, 5/6Nx SD group presented a markedly loss of renal function, increased plasma inflammatory cytokines and increased oxidative stress with a reduced activity of nitric oxide. The higher macrophage infiltration and fibrosis confirmed these conditions. RE attenuated systolic blood pressure and renal function decrease and also improved serum lipid parameters in 5/6 Nx animals. It was evident the increase of muscle strength and mass in the trained groups while the sedentary group showed reduced muscle weight and strength compared to Sham SD. CONCLUSIONS: RE implemented following 5/6Nx retard the progression of chronic kidney injury while simultaneously allowed the maintenance of skeletal muscle strength.

Resistance training attenuates inflammation and the progression of renal fibrosis in chronic renal disease.

Patients with chronic kidney disease (CKD) have progressive renal fibrosis, inflammation, and reduced muscle mass and strength. Resistance training (RT) has been suggested to mitigate the loss of muscle mass, of strength and the inflammation in CKD, but the mechanisms are unknown. The aim of this study was to evaluate the influence of RT on renal fibrosis, renal cytokine expression, creatine kinase levels, and muscle mass and strength in CKD rats. A CKD model was obtained by 5/6 nephrectomy (Nx). Fifteen 8-week-old male rats were divided into 3 groups: Sham (control), Nx SED (CKD sedentary) and Nx RT (CKD trained). The RT consisted of ladder climbing at 70% of the animal's maximal carrying capacity for 10 weeks. Muscle strength, creatine kinase levels, renal fibrosis and mRNA interleukin (IL)-4, IL-6 and IL-10 were analyzed after the RT protocol. There was significant improvement in the muscle strength and creatine kinase levels in the Nx RT group. Moreover, renal fibrosis and inflammation were attenuated, with increased IL-4 and IL-10 expression and reduced IL-6 expression in the Nx RT group compared with that in the Nx SED group. No difference in muscle mass was observed among the groups. In conclusion, RT was effective in reducing fibrosis and inflammation, in addition to increasing muscle strength and creatine kinase levels, in rats with CKD, independent of muscle mass.

TLR2 and TLR4 play opposite role in autophagy associated with cisplatin-induced acute kidney injury.

Acute kidney injury (AKI) is considered an inflammatory disease in which toll-like receptors (TLRs) signaling pathways play an important role. The activation of TLRs results in production of several inflammatory cytokines leading to further renal damage. In contrast, TLRs are key players on autophagy induction, which is associated with a protective function on cisplatin-induced AKI. Hence, the present study aimed to evaluate the specific participation of TLR2 and TLR4 molecules on the development of cisplatin-induced AKI. Complementarily, we also investigated the link between TLRs and heme oxygenase-1 (HO-1), a promisor cytoprotective molecule. First, we observed that only the absence of TLR2 but not TLR4 in mice exacerbated the renal dysfunction, tissue injury and mortality rate, even under an immunologically privileged microenvironment. Second, we demonstrated that TLR2 knockout (KO) mice presented lower expression of autophagy-associated markers when compared with TLR4 KO animals. Similar parameter was confirmed in vitro, using tubular epithelial cells derived from both KO mice. To test the cross-talking between HO-1 and TLRs, hemin (an HO-1 internal inducer) was administrated in cisplatin-treated TLR2 and TLR4 KO mice and it was detected an improvement in the global renal tissue parameters. However, this protection was less evident at TLR2 KO mice. In summary, we documented that TLR2 plays a protective role in cisplatin-induced AKI progression, in part, by a mechanism associated with autophagy up-regulation, considering that its interplay with HO-1 can promote renal tissue recover.

Intragraft transcriptional profiling of renal transplant patients with tubular dysfunction reveals mechanisms underlying graft injury and recovery.

BACKGROUND: Proximal tubular dysfunction (PTD) is associated with a decreased long-term graft survival in renal transplant patients and can be detected by the elevation of urinary tubular proteins. This study investigated transcriptional changes in biopsies from renal transplant patients with PTD to disclose molecular mechanisms underlying graft injury and functional recovery. METHODS: Thirty-three renal transplant patients with high urinary levels of retinol-binding protein, a biomarker of PTD, were enrolled in the study. The initial immunosuppressive scheme included azathioprine, cyclosporine, and steroids. After randomization, 18 patients (group 2) had their treatment modified by reducing cyclosporine dosage and substituting azathioprine for mycophenolate mofetil, while the other 15 patients (group 1) remained under the initial scheme. Patients were biopsied at enrollment and after 12 months of follow-up, and paired comparisons were performed between their intragraft gene expression profiles. The differential transcriptome profiles were analyzed by constructing gene co-expression networks and identifying enriched functions and central nodes in each network. RESULTS: Only the alternative immunosuppressive scheme used in group 2 ameliorated renal function and tubular proteinuria after 12 months of follow-up. Intragraft molecular changes observed in group 2 were linked to autophagy, extracellular matrix, and adaptive immunity. Conversely, gene expression changes in group 1 were related to fibrosis, endocytosis, ubiquitination, and endoplasmic reticulum stress. CONCLUSION: These results suggest that molecular networks associated with the control of endocytosis, autophagy, protein overload, fibrosis, and adaptive immunity may be involved in improvement of graft function.

A Regulatory miRNA-mRNA Network Is Associated with Tissue Repair Induced by Mesenchymal Stromal Cells in Acute Kidney Injury.

Mesenchymal stromal cells (MSCs) orchestrate tissue repair by releasing cell-derived microvesicles (MVs), which, presumably by small RNA species, modulate global gene expression. The knowledge of miRNA/mRNA signatures linked to a reparative status may elucidate some of the molecular events associated with MSC protection. Here, we used a model of cisplatin-induced kidney injury (acute kidney injury) to assess how MSCs or MVs could restore tissue function. MSCs and MVs presented similar protective effects, which were evidenced in vivo and in vitro by modulating apoptosis, inflammation, oxidative stress, and a set of prosurvival molecules. In addition, we observed that miRNAs (i.e., miR-880, miR-141, miR-377, and miR-21) were modulated, thereby showing active participation on regenerative process. Subsequently, we identified that MSC regulates a particular miRNA subset which mRNA targets are associated with Wnt/TGF-ß, fibrosis, and epithelial-mesenchymal transition signaling pathways. Our results suggest that MSCs release MVs that transcriptionally reprogram injured cells, thereby modulating a specific miRNA-mRNA network.

Simvastatin ameliorates experimental autoimmune encephalomyelitis by inhibiting Th1/Th17 response and cellular infiltration.

AIM: Experimental autoimmune encephalomyelitis (EAE) is a CD4(+)-mediated autoimmune pathology of the central nervous system (CNS) that is used as a model for the study of the human neuroinflammatory disease, multiple sclerosis. During the development of EAE, auto-reactive Th1 and Th17 CD4(+) T cells infiltrate the CNS promoting inflammatory cells recruitment, focal inflammation and tissue destruction. In this sense, statins, agents used to lower lipid levels, have recently shown to exert interesting immunomodulatory function. In fact, statins promote a bias towards a Th2 response, which ameliorates the clinical outcome of EAE. Additionally, simvastatin can inhibit Th17 differentiation. However, many other effects exerted on the immune system by statins have yet to be clarified, in particular during neuroinflammation. Thus, the aim of this study was to investigate the effects of simvastatin on the development of experimental autoimmune encephalomyelitis. METHODS: Mice were immunized with MOG(35-55) and EAE severity was assessed daily and scored using a clinical scale. Cytokine secretion by mononuclear cells infiltrating the CNS was evaluated by flow cytometry. RESULTS: Simvastatin (5 mg/kg/day) improved clinical outcome, induced an increase in TGF-ß mRNA expression and inhibited IL-6, IL-12p40, IL-12p70, RANTES and MIP-1ß secretion (p < 0.05). This was accompanied by a significant decrease in CNS inflammatory mononuclear cell infiltration, with reduced frequencies of both Th1 and Th17 cells. Simvastatin inhibited the proliferation of T lymphocytes co-cultured with primary microglial cells. CONCLUSIONS: Simvastatin treatment promotes EAE clinical amelioration by inhibiting T cell proliferation and CNS infiltration by pathogenic Th1 and Th17 cells.

Kinin B2 receptor does not exert renoprotective effects on mice with glycerol-induced rhabdomyolysis.

AIM: To investigate a potential protective role of the kinin B2 receptor in a glycerol-induced rhabdomyolysis mouse model. METHODS: We separated 28 C57Bl/6 male mice into 4 groups: untreated WT animals, untreated B2 knockout mice, glycerol-treated WT and glycerol-treated B2 knockout mice. Glycerol-treated animals received one intramuscular injections of glycerol solution (50% v/v, 7 mL/kg). After 48 h, urine and blood samples were collected to measure creatinine and urea levels. Additionally, kidney samples were extracted for histological evaluation, and the mRNA expression levels of kinin B1 and B2 receptors and inflammatory mediators were measured by real-time polymerase chain reaction. RESULTS: Serum creatinine and urea levels showed differences between untreated wild-type and glycerol-treated wild-type mice (0.66 ± 0.04 vs 2.61 ± 0.53 mg/dL, P < 0.01; and 33.51 ± 2.08 vs 330.2 ± 77.7 mg/dL, P < 0.005), and between untreated B2 knockout mice and glycerol-treated knockout mice (0.56 ± 0.03 vs 2.23 ± 0.87 mg/dL, P < 0.05; and 42.49 ± 3.2 vs 327.2 ± 58.4 mg/dL, P < 0.01), but there was no difference between the glycerol-treated wild-type and glycerol-treated knockout mice. Glycerol was able to induce a striking increase in kinin B2 receptor expression (> 30 times, 31.34 ± 8.9) in kidney. Animals injected with glycerol had a higher degree of tubular injury than untreated animals. Wild-type and knockout mice treated with glycerol intramuscularly present kidney injury, with impairment in renal function. However, B2 knockout mice treated with glycerol did not show a different phenotype regarding kidney injury markers, when compared to the wild-type glycerol-treated group. CONCLUSION: We conclude that the kinin B2 receptor does not have a protective role in renal injury.

Standardization of renal function evaluation in Wistar rats (Rattus norvegicus) from the Federal University of Juiz de Fora's colony.

INTRODUCTION: There is great interest in the use of animal models in the study of renal pathophysiology requires standardization of parameters. OBJECTIVE: Standardize assessment of renal function in rats from in the Center for Reproductive Biology of Federal University of Juiz de Fora's colony. METHODS: Thirty Wistar rats were used and performed measurements of creatinine (serum and urine), serum urea and proteinuria. Were evaluated: the urine collection interval in metabolic cages (24 hours or 12 hours), the need for 12-hour fast, the need of urine and serum deproteinization for creatinine measurement, need of serum deproteinization in animals with acute kidney injury to a spectrophotometer and ELISA, and the comparison of 24-hour proteinuria (PT 24 hours) with the protein/creatinine ratio (rP/C). Means were compared by the Student's t test, Pearson correlation, Bland-Altman plot for agreement and linear regression model to estimate PT 24 hours from rP/C. RESULTS: The 24 hours urine output was greater than 12 hours, interfering with the creatinine clearance calculation. In the fasting group showed less water intake and lower urinary creatinine. There was great variability for the deproteinized whey and readings performed in the two devices were similar. There was a strong correlation between PT 24 hours and rP/C and the equation was generated: PT 24 hours = (8.6113 x rP/C) + 1.0869. CONCLUSION: Was standardized: 24-hour urine collection without fasting. The deproteinization showed no benefit. The measurements were performed with spectrophotometer reliability. It generated a practical formula for estimating PT 24 hours through rP/C.

Balance between the two kinin receptors in the progression of experimental focal and segmental glomerulosclerosis in mice.

Focal and segmental glomerulosclerosis (FSGS) is one of the most important renal diseases related to end-stage renal failure. Bradykinin has been implicated in the pathogenesis of renal inflammation, whereas the role of its receptor 2 (B2RBK; also known as BDKRB2) in FSGS has not been studied. FSGS was induced in wild-type and B2RBK-knockout mice by a single intravenous injection of Adriamycin (ADM). In order to further modulate the kinin receptors, the animals were also treated with the B2RBK antagonist HOE-140 and the B1RBK antagonist DALBK. Here, we show that the blockage of B2RBK with HOE-140 protects mice from the development of FSGS, including podocyte foot process effacement and the re-establishment of slit-diaphragm-related proteins. However, B2RBK-knockout mice were not protected from FSGS. These opposite results were due to B1RBK expression. B1RBK was upregulated after the injection of ADM and this upregulation was exacerbated in B2RBK-knockout animals. Furthermore, treatment with HOE-140 downregulated the B1RBK receptor. The blockage of B1RBK in B2RBK-knockout animals promoted FSGS regression, with a less-inflammatory phenotype. These results indicate a deleterious role of both kinin receptors in an FSGS model and suggest a possible cross-talk between them in the progression of disease.

Padronização da avaliação da função renal de ratos (Rattus norvegicus) Wistar do biotério da Universidade Federal de Juiz de Fora / Standardization of renal function evaluation in Wistar rats (Rattus norvegicus) from the Federal University of Juiz de Fora's colony

Introdução: Há grande interesse na utilização de modelos animais na pesquisa da fisiopatologia renal, que requer padronização dos parâmetros analisados. Objetivo: Padronizar avaliação da função renal de ratos da colônia do biotério do Centro de Biologia da Reprodução da Universidade Federal de Juiz de Fora. Métodos: Foram utilizados 30 ratos Wistar e realizadas dosagens de creatinina (sérica e urinária), ureia sérica e proteinúria. Foram avaliados: o intervalo de coleta de urina nas gaiolas metabólicas (24 horas ou 12 horas); a necessidade de jejum de 12 horas; a necessidade de desproteinização das amostras de urina e soro para dosagens de creatinina; necessidade de desproteinização do soro de animais com injúria renal aguda (IRA) para leitura em espectrofotômetro e ELISA, além da comparação da proteinúria de 24 horas (PT 24 horas) com a relação proteína/creatinina (rP/C). Os resultados foram comparados pelos teste t de Student, correlação de Pearson, gráfico de Bland-Altman para concordância e modelo de regressão linear para estimar a PT 24 horas a partir da rP/C. Resultados: A diurese de 24 horas foi maior do que a de 12 horas, interferindo na depuração da creatinina. No grupo em jejum, houve menor ingestão hídrica e menor creatinina urinária. Houve grande variabilidade para o soro desproteinizado e as leituras realizadas nos dois equipamentos foram semelhantes. Houve forte correlação entre PT 24 horas e rP/C e foi gerada a equação: PT 24 horas = (8,6113 x rP/C) + 1,0869. Conclusão: Foi padronizada: coleta de urina em 24 horas sem jejum. A desproteinização não mostrou benefício. As dosagens foram realizadas com confiabilidade em espectrofotômetro. Foi ...

Introduction: There is great interest in the use of animal models in the study of renal pathophysiology requires standardization of parameters. Objective: Standardize assessment of renal function in rats from in the Center for Reproductive Biology of Federal University of Juiz de Fora's colony. Methods: Thirty Wistar rats were used and performed measurements of creatinine (serum and urine), serum urea and proteinuria. Were evaluated: the urine collection interval in metabolic cages (24 hours or 12 hours), the need for 12-hour fast, the need of urine and serum deproteinization for creatinine measurement, need of serum deproteinization in animals with acute kidney injury to a spectrophotometer and ELISA, and the comparison of 24-hour proteinuria (PT 24 hours) with the protein/creatinine ratio (rP/C). Means were compared by the Student's t test, Pearson correlation, Bland-Altman plot for agreement and linear regression model to estimate PT 24 hours from rP/C. Results: The 24 hours urine output was greater than 12 hours, interfering with the creatinine clearance calculation. In the fasting group showed less water intake and lower urinary creatinine. There was great variability for the deproteinized whey and readings performed in the two devices were similar. There was a strong correlation between PT 24 hours and rP/C and the equation was generated: PT 24 hours = (8.6113 x rP/C) + 1.0869. Conclusion: Was standardized: 24-hour urine collection without fasting. The deproteinization showed no benefit. The measurements were performed with spectrophotometer reliability. It generated a practical formula for estimating PT 24 hours through rP/C. .

Low-level laser therapy decreases renal interstitial fibrosis.

OBJECTIVE: the purpose of this study was to investigate the effect of low-level laser therapy (LLLT) on chronic kidney disease (CKD) in a model of unilateral ureteral obstruction (UUO). BACKGROUND DATA: Regardless of the etiology, CKD involves progressive widespread tissue fibrosis, tubular atrophy, and loss of kidney function. This process also occurs in kidney allograft. At present, effective therapies for this condition are lacking. We investigated the effects of LLLT on the interstitial fibrosis that occurs after experimental UUO in rats. METHODS: The occluded kidney of half of the 32 Wistar rats that underwent UUO received a single intraoperative dose of LLLT (AlGaAs laser, 780 nm, 22.5 J/cm(2), 30 mW, 0.75 W/cm(2), 30 sec on each of nine points). After 14 days, renal fibrosis was assessed by Sirius red staining under polarized light. Immunohistochemical analyses quantitated the renal tissue cells that expressed fibroblast (FSP-1) and myofibroblast (α-SMA) markers. Reverse transcriptase polymerase chain reaction (RT-PCR) was performed to determine the mRNA expression of interleukin (IL)-6, monocyte chemotactic protein-1 (MCP-1), transforming growth factor (TGF)-ß1 and Smad3. RESULTS: The UUO and LLLT animals had less fibrosis than the UUO animals, as well having decreased expression inflammatory and pro-fibrotic markers. CONCLUSIONS: For the first time, we showed that LLLT had a protective effect regarding renal interstitial fibrosis. It is conceivable that by attenuating inflammation, LLLT can prevent tubular activation and transdifferentiation, which are the two processes that mainly drive the renal fibrosis of the UUO model.

Transcriptome analysis of renal ischemia/reperfusion injury and its modulation by ischemic pre-conditioning or hemin treatment.

Ischemia/reperfusion injury (IRI) is a leading cause of acute renal failure. The definition of the molecular mechanisms involved in renal IRI and counter protection promoted by ischemic pre-conditioning (IPC) or Hemin treatment is an important milestone that needs to be accomplished in this research area. We examined, through an oligonucleotide microarray protocol, the renal differential transcriptome profiles of mice submitted to IRI, IPC and Hemin treatment. After identifying the profiles of differentially expressed genes observed for each comparison, we carried out functional enrichment analysis to reveal transcripts putatively involved in potential relevant biological processes and signaling pathways. The most relevant processes found in these comparisons were stress, apoptosis, cell differentiation, angiogenesis, focal adhesion, ECM-receptor interaction, ion transport, angiogenesis, mitosis and cell cycle, inflammatory response, olfactory transduction and regulation of actin cytoskeleton. In addition, the most important overrepresented pathways were MAPK, ErbB, JAK/STAT, Toll and Nod like receptors, Angiotensin II, Arachidonic acid metabolism, Wnt and coagulation cascade. Also, new insights were gained about the underlying protection mechanisms against renal IRI promoted by IPC and Hemin treatment. Venn diagram analysis allowed us to uncover common and exclusively differentially expressed genes between these two protective maneuvers, underscoring potential common and exclusive biological functions regulated in each case. In summary, IPC exclusively regulated the expression of genes belonging to stress, protein modification and apoptosis, highlighting the role of IPC in controlling exacerbated stress response. Treatment with the Hmox1 inducer Hemin, in turn, exclusively regulated the expression of genes associated with cell differentiation, metabolic pathways, cell cycle, mitosis, development, regulation of actin cytoskeleton and arachidonic acid metabolism, suggesting a pleiotropic effect for Hemin. These findings improve the biological understanding of how the kidney behaves after IRI. They also illustrate some possible underlying molecular mechanisms involved in kidney protection observed with IPC or Hemin treatment maneuvers.

Oxidative stress and modification of renal vascular permeability are associated with acute kidney injury during P. berghei ANKA infection.

Malaria associated-acute kidney injury (AKI) is associated with 45% of mortality in adult patients hospitalized with severe form of the disease. However, the causes that lead to a framework of malaria-associated AKI are still poorly characterized. Some clinical studies speculate that oxidative stress products, a characteristic of Plasmodium infection, as well as proinflammatory response induced by the parasite are involved in its pathophysiology. Therefore, we aimed to investigate the development of malaria-associated AKI during infection by P. berghei ANKA, with special attention to the role played by the inflammatory response and the involvement of oxidative stress. For that, we took advantage of an experimental model of severe malaria that showed significant changes in the renal pathophysiology to investigate the role of malaria infection in the renal microvascular permeability and tissue injury. Therefore, BALB/c mice were infected with P. berghei ANKA. To assess renal function, creatinine, blood urea nitrogen, and ratio of proteinuria and creatininuria were evaluated. The products of oxidative stress, as well as cytokine profile were quantified in plasma and renal tissue. The change of renal microvascular permeability, tissue hypoxia and cellular apoptosis were also evaluated. Parasite infection resulted in renal dysfunction. Furthermore, we observed increased expression of adhesion molecule, proinflammatory cytokines and products of oxidative stress, associated with a decrease mRNA expression of HO-1 in kidney tissue of infected mice. The measurement of lipoprotein oxidizability also showed a significant increase in plasma of infected animals. Together, our findings support the idea that products of oxidative stress, as well as the immune response against the parasite are crucial to changes in kidney architecture and microvascular endothelial permeability of BALB/c mice infected with P. berghei ANKA.

MyD88 signaling pathway is involved in renal fibrosis by favoring a TH2 immune response and activating alternative M2 macrophages.

Inflammation contributes to the pathogenesis of chronic kidney disease (CKD). Molecules released by the inflamed injured tissue can activate toll-like receptors (TLRs), thereby modulating macrophage and CD4(+) T-cell activity. We propose that in renal fibrogenesis, M2 macrophages are recruited and activated in a T helper subset 2 cell (T(H)2)-prone inflammatory milieu in a MyD88-dependent manner. Mice submitted to unilateral ureteral ligation (UUO) demonstrated an increase in macrophage infiltration with collagen deposition after 7 d. Conversely, TLR2, TLR4 and MyD88 knockout (KO) mice had an improved renal function together with diminished T(H)2 cytokine production and decreased fibrosis formation. Moreover, TLR2, TLR4 and MyD88 KO animals exhibited less M2 macrophage infiltration, namely interleukin (IL)-10(+) and CD206(+) CD11b(high) cells, at 7 d after surgery. We evaluated the role of a T(H)2 cytokine in this context, and observed that the absence of IL-4 was associated with better renal function, decreased IL-13 and TGF-ß levels, reduced arginase activity and a decrease in fibrosis formation when compared with IL-12 KO and wild-type (WT) animals. Indeed, the better renal outcomes and the decreased fibrosis formation were restricted to the deficiency of IL-4 in the hematopoietic compartment. Finally, macrophage depletion, rather than the absence of T cells, led to reduced lesions of the glomerular filtration barrier and decreased collagen deposition. These results provide evidence that future therapeutic strategies against renal fibrosis should be accompanied by the modulation of the M1:M2 and T(H)1:T(H)2 balance, as T(H)2 and M2 cells are predictive of fibrosis toward mechanisms that are sensed by innate immune response and triggered in a MyD88-dependent pathway.

TLR2, TLR4 and the MYD88 signaling pathway are crucial for neutrophil migration in acute kidney injury induced by sepsis.

The aim of this study was to investigate the role of TLR2, TLR4 and MyD88 in sepsis-induced AKI. C57BL/6 TLR2(-/-), TLR4(-/-) and MyD88(-/-) male mice were subjected to sepsis by cecal ligation and puncture (CLP). Twenty four hours later, kidney tissue and blood samples were collected for analysis. The TLR2(-/-), TLR4(-/-) and MyD88(-/-) mice that were subjected to CLP had preserved renal morphology, and fewer areas of hypoxia and apoptosis compared with the wild-type C57BL/6 mice (WT). MyD88(-/-) mice were completely protected compared with the WT mice. We also observed reduced expression of proinflammatory cytokines in the kidneys of the knockout mice compared with those of the WT mice and subsequent inhibition of increased vascular permeability in the kidneys of the knockout mice. The WT mice had increased GR1(+low) cells migration compared with the knockout mice and decreased in GR1(+high) cells migration into the peritoneal cavity. The TLR2(-/-), TLR4(-/-), and MyD88(-/-) mice had lower neutrophil infiltration in the kidneys. Depletion of neutrophils in the WT mice led to protection of renal function and less inflammation in the kidneys of these mice. Innate immunity participates in polymicrobial sepsis-induced AKI, mainly through the MyD88 pathway, by leading to an increased migration of neutrophils to the kidney, increased production of proinflammatory cytokines, vascular permeability, hypoxia and apoptosis of tubular cells.

The polysaccharide fraction of Propionibacterium acnes modulates the development of experimental focal segmental glomerulosclerosis.

The pathogenesis of focal segmental glomerulosclerosis (FSGS) appears to be associated with type-2 cytokines and podocyte dysfunction. In this study, we tested the hypothesis that immunization with the polysaccharide fraction of Propionibacterium acnes (PS), a pro-Th1 agonist, may subvert the type-2 profile and protect podocytes from adriamycin-induced glomerulosclerosis. Adriamycin injection resulted in albuminuria and increased serum creatinine in association with loss of glomerular podocin and podoplanin expression, which is consistent with podocyte dysfunction. Renal tissue analysis revealed the expression of transcripts for GATA3 and fibrogenic-related proteins, such as TGF-ß, tissue inhibitor of metalloproteinase-1 (TIMP-1) and metalloproteinase 9 (MMP9). In association with the expression of fibrogenic transcripts, we observed peri-glomerular expression of α-smooth muscle actin (α-SMA), indicating epithelial-to-mesenchymal transition, and increased expression of proliferating cell nuclear antigen (PCNA) in tubular cells, suggesting intense proliferative activity. Previous immunization with PS inhibited albuminuria and serum creatinine in association with the preservation of podocyte proteins and inhibition of fibrogenic transcripts and the expression of α-SMA and PCNA proteins. Tissue analysis also revealed that PS treatment induced expression of mRNA for GD3 synthase, which is a glycosiltransferase related to the synthesis of GD3, a ganglioside associated with podocyte physiology. In addition, PS treatment inhibited the influx of inflammatory CD8(pos) and CD11b(pos) cells to kidney tissue. Finally, PS treatment on day 4 post-ADM, a period when proteinuria was already established, was able to improve renal function. Thus, we demonstrate that the PS fraction of P. acnes can inhibit FSGS pathogenesis, suggesting that immunomodulation can represent an alternative approach for disease management.