Errol D. Crook

Transcriptional Regulation of Transforming Growth Factor β1 by Glucose: Investigation into the Role of the Hexosamine Biosynthesis Pathway

BACKGROUND The hexosamine biosynthesis pathway (HBP) is hypothesized to mediate many of the adverse effects of hyperglycemia. We have shown previously that increased flux through this pathway leads to induction of the growth factor transforming growth factor-alpha (TGF-alpha) and to insulin resistance in cultured cells and transgenic mice. TGF-beta is regulated by glucose and is involved in the development of diabetic nephropathy. We therefore hypothesized that the HBP was involved in the regulation of TGF-beta by glucose in rat vascular and kidney cells. METHODS A plasmid containing the promoter region of TGF-beta1 cloned upstream of the firefly luciferase gene was electroporated into rat aortic smooth muscle, mesangial, and proximal tubule cells. Luciferase activity was measured in cellular extracts from cells cultured in varying concentrations of glucose and glucosamine. RESULTS Glucose treatment of all cultured cells led to a time- and dose-dependent stimulation in TGF-beta1 transcriptional activity, with high (20 mM) glucose causing a 1.4- to 2.0-fold increase. Glucose stimulation did not occur until after 12 hours and disappeared after 72 hours of treatment. Glucosamine was more potent than glucose, with 3 mM stimulating up to a 4-fold increase in TGFbeta1-transcriptional activity. The stimulatory effect of glucosamine was also dose-dependent but was slower to develop and longer lasting than that of glucose. CONCLUSIONS The metabolism of glucose through the HBP mediates extracellular matrix production, possibly via the stimulation of TGF-beta in kidney cells. Hexosamine metabolism therefore, may play a role in the development of diabetic nephropathy.

Glucose-induced insulin resistance of phosphatidylinositol 3′-OH kinase and AKT/PKB is mediated by the hexosamine biosynthesis pathway

Hyperglycemia is responsible for many of the vascular complications and metabolic derangements seen in diabetes. One potential regulator of the effects of glucose is the hexosamine biosynthesis pathway (HBP). Glutamine: fructose-6-phosphate amidotransferase (GFA), the first and rate-limiting enzyme in this pathway, catalyzes the transfer of an amino group from glutamine to fructose-6-phosphate to form glucosamine-6-phosphate. Overexpression of GFA in rat-1 fibroblasts results in insulin resistance for glycogen synthase (GS) activity, and renders these cells more sensitive to the effects of glucose. Using rat-1 cells, we examine further the mechanisms whereby hexosamines lead to insulin resistance. Insulin stimulated GS activity was found to occur via a PI-3 kinase (PI-3K)-dependent pathway as wortmannin, an inhibitor of PI-3K, blocked insulins ability to stimulate GS activity. Subsequently, we examined the effects of hexosamines on PI-3K and Akt/PKB activity. Cells were cultured in 1 mM glucose (low glucose, LG), 20 mM glucose (high glucose, HG), or 1 mM glucose plus 3 mM glucosamine (GlcN) for 16--20 h. After treatment with insulin (100 nM) for 5 min, cell extracts were assayed for IRS-1 associated and total PI-3K activity. At LG, insulin increased PI-3K activity by 43%. There was no insulin stimulation of PI-3K activity in cells cultured in HG or GlcN. There was a trend for IRS-1 protein levels to decrease in HG but not GlcN. PI-3K protein levels were not altered by HG or GlcN. Finally PKB activity was assayed. At LG, insulin stimulated PKB activity. Again, both HG and GlcN significantly reduced insulins ability to stimulate PKB activity. We conclude that the hexosamine-mediated insulin resistance of GS activity seen in rat-1 cells is mediated by hexosamine regulation of PI-3K and PKB.

Focal Segmental Glomerulosclerosis in African Americans: Effects of Steroids and Angiotensin Converting Enzyme Inhibitors

BACKGROUND Focal segmental glomerulosclerosis (FSGS) is a common primary glomerulopathy in African Americans. In this report, we present data on 40 African American patients with FSGS from our medical center. METHODS Patients were identified from a review of all charts seen in our conservative management renal clinic in 1996, a review of renal biopsy rolls (1994-1998), and a review of patients entering the end-stage renal disease (ESRD) program with a primary diagnosis of FSGS (1993- 1997). Charts were reviewed for demographic, biopsy, and treatment data. Patients who were observed for at least 4 months (range, 4-125 months) were included. ESRD was used as the primary endpoint (n = 12). Data were analyzed using univariate and multivariate Cox hazards and Kaplan-Meier survival analysis. Twenty-four patients were treated with angiotensin-converting enzyme (ACE) inhibitors. Similarly, 24 patients were treated with corticosteroids for a mean of 8.75 +/- 2.6 months and a total dose of 9.3 +/- 2.2 g. RESULTS On univariate analysis, factors found to be significant determinants for reaching ESRD were the initial creatinine (P = 0.0001), interstitial fibrosis (P = 0.032), the percentage of globally sclerosed glomeruli (P = 0.0018), and the mean arterial blood pressure over the course of follow-up (P = 0.05). Neither the ACE inhibitors nor the corticosteroids had a significant impact on reaching ESRD. The patients reaching ESRD (n = 12) were analyzed separately. The mean time from biopsy to ESRD was 24.7 +/- 9.8 months. ACE inhibitors prolonged renal survival (P = 0.023), but steroids did not. Initial creatinine was the only factor found to be a significant determinant for ESRD. CONCLUSIONS We conclude that FSGS is common in African Americans. Early diagnosis and blood pressure control are important, but the beneficial effects of steroids and ACE inhibitors in this population are still unclear.

The Role of Hypertension, Obesity, and Diabetes in Causing Renal Vascular Disease

The Jackson Heart Study will be an epidemiological study of African Americans in Jackson, Mississippi, to identify risk factors for development and progression of cardiovascular disease. One of the potential risk factors to be assessed in this study is renal vascular disease. Atherosclerotic renal vascular disease is a disease of the elderly, is predominantly seen in white people, and is strongly associated with diffuse atherosclerotic disease and high-grade hypertensive retinopathy. Patients with ischemic nephropathy may constitute up to 16% of new dialysis patients and die more quickly while on renal replacement therapy. Although often not present, hypertension is a commonly observed consequence (but probably not a cause) of renal vascular disease, and the control of blood pressure may not halt the progression of the disease. Approximately 20-25% of patients with moderate to severe renal artery stenosis will be diabetic. Diabetic patients fair less well with intervention and have a higher progression to end-stage renal disease or death. Obesity is not commonly seen in patients with renal vascular disease. The Jackson Heart Study may be able to assess the true incidence of atherosclerotic renal vascular disease in African Americans and its impact of cardiovascular morbidity and mortality.

Diabetic nephropathy in African Americans

Diabetic nephropathy (DN) is the number one cause of end-stage renal disease in United States and is highly prevalent in African Americans. We have found that among African Americans in Mississippi diabetic nephropathy appears to affect females more than males, which may be related to increased rates of obesity and diabetes in African American women. Glycemic control and control of blood pressure is essential to prolong renal survival and to protect against cardiovascular events. Angiotensin-converting enzyme inhibitors reduce cardiovascular mortality in diabetics and are tolerated in advanced renal disease. The impact of glycemic control, appropriate antihypertensives, and the optimal level of blood pressure control in African Americans with advanced DN require further study. This article reviews the impact, clinical characteristics, risk factors, and treatment of diabetic nephropathy in African Americans.

Caring for Adolescent Family Members of Physician Colleagues

It is recommended that physicians do not care for their family members. However, there are also many concerning issues when physicians care for the family members of their colleagues. This can be particularly challenging when the patient is an adolescent. We present the case of Dr. B who is asked to see the teenage daughter of her colleague Dr. A, and discuss the issues of caring for an adolescent family member of a physician colleague. Patient confidentiality, autonomy, and maintenance of the patient-physician relationship are core principles that must be adhered to in this situation. The roles of the treating physician and the parental physician are discussed.

Renal Scars Masquerading As Complex Masses in a Patient with Vesicoureteral Reflux Nephropathy

Vesicoureteral reflux can lead to chronic pyelonephritis, renal scarring, and renal failure. We present a case of renal scarring masquerading as bilateral, complex renal masses. A 35-year old woman who was diagnosed with vesicoureteral reflux as a child presented for evaluation of recently developed hypertension and an abnormal renal ultrasound. Her serum creatinine level was 2.5 mg/dL and she had subnephrotic-range proteinuria. A renal sonogram showed small, echogenic kidneys and bilateral complex renal masses of 3.8 (right) and 4.4 (left) cm in greatest dimensions. CT scan of the kidneys revealed slightly contrast-enhancing masses with irregular walls. Renal angiogram showed decreased blood supply to the areas coinciding with the masses consistent with renal scarring. There was no increased vascularity. This case demonstrates that renal scarring may masquerade as renal masses. A step-wise, comprehensive approach is necessary to rule out potentially malignant lesions in these patients.