Regulation of indoleamine 2,3 dioxygenase and its role in a porcine model of acute kidney allograft rejection.

In kidney transplantation acute allograft rejection is the most common cause of late allograft loss. Changes in indoleamine 2,3 dioxygenase (IDO) activity, which catabolizes the degradation of tryptophan to kynurenine, may predict rejection. However, exogenous IDO is immunosuppressive in rodent kidney transplantation. Thus, the increase in IDO activity observed in acute allograft rejection is insufficient to prevent rejection. To address this question, we assessed the regulation of IDO and its role in acute rejection in a porcine model of kidney transplant. In tissue samples from rejecting kidney allografts, we showed a 13-fold increase in IDO gene transcription and 20-fold increase in IDO enzyme activity when compared with autotransplanted kidneys. Allografts also demonstrated an over fourfold increase in tissue interferon (IFN)-γ, with marked increases in tumor necrosis factor (TNF)-α, TNF-ß and interleukin 1ß. Gene transcription and protein levels of kynurenine 3-monooxygenase (KMO) were decreased. KMO generates the immunosuppressive kynurenine, 3-hydroxykynurenine. The results of these studies demonstrate a clear association between rejection and increased allograft IDO expression, likely driven in part by IFN-γ and facilitated by other cytokines of the allogeneic response. Moreover, the loss of downstream enzymatic activity in the IDO metabolic pathway may suggest novel mechanisms for the perpetuation of rejection.

Autophagy is activated to protect against podocyte injury in adriamycin-induced nephropathy.

Podocytes are highly differentiated epithelial cells wrapping glomerular capillaries to form the filtration barrier in kidneys. As such, podocyte injury or dysfunction is a critical pathogenic event in glomerular disease. Autophagy plays an important role in the maintenance of the homeostasis and function of podocytes. However, it is less clear whether and how autophagy contributes to podocyte injury in glomerular disease. Here, we have examined the role of autophagy in adriamycin-induced nephropathy, a classic model of glomerular disease. We show that autophagy was induced by adriamycin in cultured podocytes in vitro and in podocytes in mice. In cultured podocytes, activation of autophagy with rapamycin led to the suppression of adriamycin-induced apoptosis, whereas inhibition of autophagy with chloroquine enhanced podocyte apoptosis during adriamycin treatment. To determine the role of autophagy in vivo, we established an inducible podocyte-specific autophagy-related gene 7 knockout mouse model (Podo-Atg7-KO). Compared with wild-type littermates, Podo-Atg7-KO mice showed higher levels of podocyte injury, glomerulopathy, and proteinuria during adriamycin treatment. Together, these observations support an important role of autophagy in protecting podocytes under the pathological conditions of glomerular disease, suggesting the therapeutic potential of autophagy induction.

GLUT1 regulation of the pro-sclerotic mediators of diabetic nephropathy.

Diabetic glomerulosclerosis is characterized by accumulation of extracellular matrix proteins, mesangial expansion, and tubulointerstitial fibrosis. Hyperglycemia accelerates development of the disease, a direct result of increased intracellular glucose availability. The facilitative glucose transporter GLUT1 mediates mesangial cell glucose flux which leads to activation of signaling cascades favoring glomerulosclerosis, including pathways mediated by angiotensin II (Ang II), transforming growth factor ß (TGF-ß), connective tissue growth factor (CTGF), and vascular endothelial growth factor (VEGF). Ang II has both hemodynamic and metabolic effects directly inducing GLUT1 and/or matrix protein synthesis through diacyl glycerol (DAG) or protein kinase C (PKC) induction, mesangial cell stretch, and/or through transactivation of the epidermal growth factor receptor, the platelet-derived growth factor receptor, and the insulin-like growth factor-1 receptor, all of which may stimulate GLUT1 synthesis via an ERK-mediated pathway. Conversely, inhibition of Ang II effects suppresses GLUT1 and cellular glucose uptake. GLUT1-mediated glucose flux leads to metabolism of glucose via glycolysis, with induction of DAG, PKC, TGF-ß1, CTGF and VEGF. VEGF in turn triggers both GLUT1 and matrix synthesis. New roles for GLUT1-mTOR and GLUT1-mechano-growth factor interactions in diabetic glomerulosclerosis have also recently been suggested. Recent mouse models confirmed roles for GLUT1 in vivo in stimulating glomerular growth factor expression, growth factor receptors and development of glomerulosclerosis. GLUT1 may therefore act in concert with cytokines and growth factors to induce diabetic glomerulosclerosis. Further clarification of the pathways involved may prove useful for the therapy of diabetic nephropathy. New directions for investigation are discussed.

Role of gastroesophageal reflux symptoms in exacerbations of COPD.

BACKGROUND AND AIMS: The impact of gastroesophageal reflux disease (GERD) on exacerbations of COPD has never been evaluated. The aims of this investigation were to determine the prevalence of gastroesophageal reflux (GER) symptoms in COPD patients and the effect of GER on the rate of exacerbations of COPD per year. METHODS: A questionnaire-based, cross sectional survey was performed. Subjects were recruited from the outpatient pulmonary clinics at the University of Florida Health Science Center/Jacksonville. Included patients had an established diagnosis of COPD. Exclusion criteria were respiratory disorders other than COPD, known esophageal disease, active peptic ulcer disease, Zollinger-Ellison syndrome, mastocytosis, scleroderma, and current alcohol abuse. Those meeting criteria and agreeing to participate were asked to complete the Mayo Clinic GERD questionnaire by either personal/telephone interview. Clinically significant reflux was defined as heartburn and/or acid regurgitation weekly. Other outcome measures noted were frequency and type of COPD exacerbations. Statistical analysis was performed using the Fisher exact test for categorical data and the independent t test for interval data. RESULTS: Eighty-six patients were enrolled and interviewed (mean age, 67.5 years). Male patients accounted for 55% of the study group. Overall, 37% of patients reported GER symptoms. The mean FEV(1) percentage of predicted was similar in those with or without GER. The rate of exacerbations of COPD was twice as high in patients with GER symptoms compared to those without GER symptoms (3.2/yr vs 1.6/yr, p = 0.02). CONCLUSIONS: The presence of GER symptoms appears to be associated with increased exacerbations of COPD.

Proposed pathogenesis of idiopathic loin pain-hematuria syndrome.

BACKGROUND: To study loin pain-hematuria syndrome (LPHS) pathogenesis, we evaluated 43 consecutive patients for whom urological evaluation failed to disclose the cause of their recurrent flank pain and hematuria. Each underwent percutaneous kidney biopsy. In 9 patients, the biopsy specimen showed immunoglobulin A nephritis, an established cause of LPHS. We suggest these cases be designated secondary LPHS. They are not included in this analysis. The remaining patients (N = 34) are designated idiopathic (primary) LPHS. They are the basis of this report. METHODS: Demographics of patients with primary LPHS are mean age of 30.8 +/- 10.3 years; 74% women; 94% white; and history of kidney stones, 47%, although none was obstructing. RESULTS: Primary LPHS kidney biopsy specimens showed red blood cells (RBCs) in multiple tubules, consistent with glomerular hematuria. Glomeruli were normal by means of light and immunofluorescent microscopy; however, more than 50% of biopsy specimens showed unusually thin or thick glomerular basement membranes. To assess whether the biopsy itself caused RBCs in tubules, we compared RBCs in renal tubular cross-sections from primary LPHS biopsies with those of normal kidneys (donors, n = 10). The mean percentage of tubular cross-sections containing RBCs was greater in primary LPHS than normal specimens (7.2% +/- 6.5% versus 1.6% +/- 1.0% [SD]; P < 0.0001), confirming glomerular hematuria in patients with primary LPHS. CONCLUSION: Primary LPHS pathogenesis includes glomerular hematuria, apparently from structurally abnormal glomerular basement membrane. Primary LPHS pain may be initiated by obstructing RBC casts and perhaps microcrystals in those with a history of urolithiasis. Nevertheless, other factors are needed to explain the severe pain in patients with primary LPHS.

Reactive oxygen species and EGR-1 gene expression in surgical postoperative peritoneal adhesions.

Postoperative peritoneal adhesions are common and serious complications of general abdominal and gynecological surgery that can lead to chronic abdominal pain, small-bowel obstruction and infertility. The specific pathophysiology of peritoneal adhesions remains elusive and current treatment is relegated to prevention through meticulous surgical technique and protective physical barriers, gels and solutions. We have reported that reactive oxygen species (ROS), generated by phagocytic cells at the site of tissue injury, serve as major signaling molecules regulating the expression of vascular endothelial growth factor (VEGF) and subsequent wound repair. We hypothesized that peritoneal adhesions are a product of over-healing surgical wounds and that, like in wound healing, ROS are implicated in their pathogenesis. We examined the presence of footprints of ROS and the ROS-inducible angiogenic factor VEGF in human adhesion tissue. An experimental model of peritoneal adhesion was established in rodents to study of the dynamics of ROS-induced gene expression during de novo adhesion tissue formation. Immunohistochemical analysis demonstrated presence of ROS/oxidant and macrophages in human peritoneal tissue. The presence of ROS and ROS-sensitive transcription factor EGR-1 was also evident in an experimental rodent peritoneal adhesion model. Along with ROS, VEGF, and a large number of mature and immature CD31/vWF positive blood vessels were present in the adhesion tissue. These observations are not consistent with the contention that adhesions are non-functional scar tissue. The newly developed rodent model of adhesion may present a useful approach to reproducibly and objectively study molecular mechanisms underlying the dynamic process of de novo adhesion tissue formation.

Glomerular beta-galactosidase expression following transduction with microsphere-adenoviral complexes.

The aortic injection of adenoviral-microsphere complexes is a useful technique for in vivo gene transfer (transduction) to the glomerulus. In this approach, the appearance of the foreign transprotein in the glomerulus may result from glomerular cell gene transfer and local synthesis or hepatic cell transduction followed by synthesis, secretion, and deposition in the glomerulus. We postulated that glomerular expression of the foreign transgene was the result of glomerular cell transduction. To test this question, male SD rats underwent aortic injections with adenovirus containing the LacZ expression cassette [expressing beta-galactosidase (betagal)] coupled to 16 microm diameter microspheres. After 48 hours, histologic staining confirmed glomerular expression of the betagal transprotein and reverse transcription in situ polymerase chain reaction demonstrated the presence of the betagal transgene in the glomerulus. In addition, hepatic expression of the betagal transprotein was minimal and substantially less than that observed in the glomeruli. These data support the contention that adenoviral-microsphere complexes result in glomerular cell transduction with the desired transgene, followed by local transprotein synthesis. This approach may prove useful for facilitating glomerular gene transfer in the development of gene therapy for glomerulonephritis.

Fatal group a streptococcal necrotizing myopathy.

A 72-year-old woman on immunosuppressive therapy for renal transplantation was admitted to the hospital for diffuse, progressive weakness that developed over several weeks. Serum creatine kinase steadily increased and she developed myoglobinuria. Blood cultures were positive for group A Streptococcus pyogenes. The muscle biopsy demonstrated a necrotizing myopathy without cellular infiltrate. Despite treatment, she evolved to multiorgan system failure and cardiovascular collapse. This is a novel presentation of group A streptococcal muscle infection with diffuse, generalized muscle weakness. It stands in striking contrast to the usual infection with this organism confined to one extremity and associated with a prolific cellular response in the muscle.