Biomedical subjects

Richard M Green

Publications and source records attributed to Richard M Green.

At least 19 recordsLinked to original sources

PubMed2006

Subcellular compartmentalization of glutathione: correlations with parameters of oxidative stress related to genotoxicity.

Glutathione (GSH) is a major component of the antioxidant defence system of mammalian cells and is found in subcellular pools within the cytoplasm, nucleus and mitochondria. To evaluate the relationships between these pools and parameters of oxidative stress related to genotoxicity, wild type (WT) and 8-oxo-2'-deoxyguanosine glycosylase 1 (OGG1)-null (mOGG1(-/-)) mouse embryonic fibroblasts (MEF) were treated with buthionine sulphoximine (BSO; 0-1000 microM, 24 h), an inhibitor of GSH biosynthesis. BSO treatment resulted in a concentration-dependent depletion of GSH from the cytoplasm, but depletion of mitochondrial and nuclear GSH occurred only at concentrations > or =100 microM. GSH levels were correlated with reactive oxygen species (ROS), lipid peroxidation (measured as the increase in the genotoxic end-product malondialdehyde (MDA)) and oxidative DNA modifications, measured as both frank DNA strand-breaks (FSB) and oxidized purine lesions (OxP) using the alkaline comet assay with formamidopyrimidine DNA glycosylase (FPG) modification; this system allowed for the identification of BSO-induced DNA modifications as primarily mutagenic 8-oxo-2'-deoxyguanosine lesions. A number of significant correlations were observed. First, negative linear correlations were observed between mitochondrial GSH and ROS (r = -0.985 and r = -0.961 for WT and mOGG1(-/-) MEF, respectively), and mitochondrial GSH and MDA (r = -0.967 and r = -0.963 for WT and mOGG1(-/-) MEF, respectively). Second, positive linear correlations were observed between ROS and MDA (r = 0.996 and r = 0.935 for WT and mOGG1(-/-) MEF, respectively), and ROS and OxP (r = 0.938 and r = 0.981 for WT and mOGG1(-/-) MEF, respectively). Finally, oxidative DNA modifications displayed a negative linear correlation with nuclear GSH (r = -0.963 and -0.951 between nuclear GSH and FSB and OxP, respectively, for WT MEF and r = -0.960 between nuclear GSH and OxP in mOGG1(-/-) MEF), thus, demonstrating the genotoxic potential of compounds that deplete GSH. The findings highlight the critical roles of the mitochondrial and nuclear GSH pools in protecting cellular components, particularly DNA, from oxidative modification.

Animals↗

PubMed2006

Endovascular versus open mesenteric revascularization: immediate benefits do not equate with short-term functional outcomes.

BACKGROUND: Percutaneous therapy for symptomatic visceral occlusive disease is rapidly gaining popularity in many centers. This study evaluates the anatomic and functional outcomes of open and endovascular therapy for chronic mesenteric ischemia at an academic medical center. STUDY DESIGN: We performed a retrospective review of patients who underwent endovascular or open mesenteric arterial revascularization for chronic mesenteric ischemia between January 1989 and September 2003. Indications for revascularization included postprandial abdominal pain (92%) or weight loss (54%). All had atherosclerotic visceral occlusive disease with a median of 2 vessels with more than 50% stenosis or occlusion on angiography. Sixty patients (44 women, mean age 66 years) underwent 67 interventions (43 vessels bypassed, 23 vessel endarterectomies, 22 vessel angioplasty and stents). The median numbers of vessels revascularized were two in the open group and one in the endovascular group. RESULTS: Thirty-day mortality and cumulative survival at 3 years were similar (open, 15% and 62% +/- 9%; endovascular, 21% and 63%+/-14%, respectively; p=NS). Cumulative patencies at 6 months were 83%+/-7% and 68%+/-14% in the open and endovascular groups, respectively (p=NS). Major morbidity, median postoperative length of stay, and cumulative freedom from recurrent symptoms at 6 months were significantly greater in the open group (open, 46%, 23 days, and 71%+/-7%, respectively; endovascular, 19%, 1 day, and 34%+/-10%, respectively; p < 0.01). CONCLUSIONS: Endovascular revascularization is attractive because it carries equivalent patency to open revascularization. Symptomatic benefit of endovascular revascularization is not achieved, probably as a result of incomplete revascularization. Despite incomplete revascularization, endovascular therapy has equivalent survival and lower morbidity compared with open revascularization. Complete endovascular revascularization needs further evaluation to determine if it is superior to open revascularization. In the interim, endovascular therapy should be reserved for the patient unable to undergo open revascularization.

Adult↗

PubMed2006

Quantitative trait loci analysis of mice administered the methionine-choline deficient dietary model of experimental steatohepatitis.

BACKGROUND: Nonalcoholic steatohepatitis (NASH) is a common disease with a poorly understood etiology, and the methionine-choline-deficient (MCD) diet is a nutritional model of NASH. Quantitative trait loci (QTL) analysis is a standard method for chromosomal mapping of polygenic disease traits. The purpose of this study is to administer mice an MCD diet in order to determine the strain-specific susceptibility for developing steatohepatitis, and to apply a computational methodology of QTL analysis to identify associated chromosomal susceptibility loci. METHODS: Inbred mice were fed an MCD diet and alanine aminotransferase (ALT), hepatic triglycerides, liver weight, and weight loss were measured as phenotypic markers of steatohepatitis. RESULTS: A/J mice developed the highest ALT and hepatic triglyceride levels. Using linear regression analysis, gene loci affecting serum ALT levels were identified on four chromosomes, and four loci that affect liver weight were also identified. In contrast, no QTLs for hepatic triglycerides or body weight were identified. Of note, loci for ALT and liver weight co-localized to proximal segments of chromosomes 2 and 15, in regions previously identified as QTLs for liver fibrosis. CONCLUSIONS: These data indicate that experimental steatohepatitis is a polygenic disease with genes determining ALT, liver weight, and liver fibrosis.

Animals↗

PubMed2005

Mice overexpressing hepatic Abcb11 rapidly develop cholesterol gallstones.

Cholelithiasis is a polygenic disease, although the genes responsible for gallstone formation have not yet been clearly identified. QTL analysis has identified the Lith 1 loci on mouse Chromosome 2, and the hepatic bile salt transporter Abcb11 maps to the Lith 1 locus. We have used recently developed TTR-Abcb11 transgenic mice that overexpress Abcb11 to determine the effects of Abcb11 overexpression on cholesterol gallstone formation. TTR-Abcb11 and FVB/NJ strain control mice were fed a lithogenic or chow diet and cholesterol crystal and gallstone formation were measured. Biliary lipids in gallbladder bile and gene expression of canalicular lipid transporters were also analyzed. TTR-Abcb11 mice fed a lithogenic diet had an increased rate of cholesterol crystal and gallstone formation. This was associated with an increase in both the hydrophobic bile salt and cholesterol content of gallbladder bile. Expression of Abcb4, Abcg5, and Abcg8 did not change before gallstone formation. These data indicate that hepatic overexpression of Abcb11 increases the rate of cholesterol gallstone formation. This is likely because of increases in bile salt hydrophobicity but not because of alterations of other biliary lipid transporters. These findings strongly support Abcb11 as a Lith 1 gene.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2005

Mice heterozygous for the Mdr2 gene demonstrate decreased PEMT activity and diminished steatohepatitis on the MCD diet.

BACKGROUND/AIMS: The administration of a methionine and choline deficient (MCD) diet to mice serves as an animal model of NASH. The multidrug resistant 2 (Mdr2) P-glycoprotein encodes for the canalicular phospholipid transporter, and Mdr2 (+/-) mice secrete 40% less phosphatidylcholine than wild-type mice. We have hypothesized that phosphatidylethanolamine-N-methyl transferase (PEMT) up-regulation is a consequence of MCD diet administration, and is important for the pathogenesis of steatohepatitis in this model. However, the effect of decreased phosphatidylcholine secretion and modulation of PEMT on the development of diet-induced steatohepatitis in Mdr2 (+/-) mice has not been explored. Thus, the purpose of the study is to examine the effects of the MCD diet on Mdr2 (+/-) mice. METHODS: Mdr2 (+/-) and Mdr2 (+/+) mice were treated with an MCD or control diet for up to 30 days, and the severity of steatohepatitis, PEMT activity and hepatic S-adenosylmethionine (SAM), and S-adenosylhomocysteine (SAH) levels were measured. RESULTS: Serum ALT levels, hepatic inflammation, and PEMT activity were significantly lower, and hepatic SAM:SAH ratios were significantly higher in Mdr2 (+/-) mice at 7 and 30 days on the MCD diet. CONCLUSIONS: Mdr2 (+/-) mice have diminished susceptibility to MCD diet-induced NASH, which is associated with a relative decrease in PEMT activity and increased SAM:SAH ratios.

ATP Binding Cassette Transporter, Subfamily B↗

PubMed2005

Structural and mechanistic studies of Escherichia coli nitroreductase with the antibiotic nitrofurazone. Reversed binding orientations in different redox states of the enzyme.

The antibiotics nitrofurazone and nitrofurantoin are used in the treatment of genitourinary infections and as topical antibacterial agents. Their action is dependent upon activation by bacterial nitroreductase flavoproteins, including the Escherichia coli nitroreductase (NTR). Here we show that the products of reduction of these antibiotics by NTR are the hydroxylamine derivatives. We show that the reduction of nitrosoaromatics is enzyme-catalyzed, with a specificity constant approximately 10,000-fold greater than that of the starting nitro compounds. This suggests that the reduction of nitro groups proceeds through two successive, enzyme-mediated reactions and explains why the nitroso intermediates are not observed. The global reaction rate for nitrofurazone determined in this study is over 10-fold higher than that previously reported, suggesting that the enzyme is much more active toward nitroaromatics than previously estimated. Surprisingly, in the crystal structure of the oxidized NTR-nitrofurazone complex, nitrofurazone is oriented with its amide group, rather than the nitro group to be reduced, positioned over the reactive N5 of the FMN cofactor. Free acetate, which acts as a competitive inhibitor with respect to NADH, binds in a similar orientation. We infer that the orientation of bound nitrofurazone depends upon the redox state of the enzyme. We propose that the charge distribution on the FMN rings, which alters upon reduction, is an important determinant of substrate binding and reactivity in flavoproteins with broad substrate specificity.

Acetates↗

PubMed2005

Steatohepatitis develops rapidly in transgenic mice overexpressing Abcb11 and fed a methionine-choline-deficient diet.

Nonalcoholic fatty liver disease is the most common reason for abnormal liver chemistries in the United States. The factors that lead from benign steatosis to nonalcoholic steatohepatitis are poorly understood. Transthyretin-Abcb11 (TTR-Abcb11) transgenic mice overexpress the bile salt transporter Abcb11 and hypersecrete biliary lipids. Thus the aim of this study is to employ feeding of the methionine-choline-deficient (MCD) diet to TTR-Abcb11 transgenic mice to further determine the mechanisms responsible for the development of steatohepatitis. FVB/NJ and TTR-Abcb11 mice were fed control or MCD diets for up to 30 days. Serum aminotransferase levels, serum and hepatic triglyceride content, cytokines, markers of oxidative stress, and expression of selective genes were examined. MCD diet-fed TTR-Abcb11, but not wild-type, mice have elevated serum aminotransferase levels when compared after 7 days. They also have significantly lower hepatic triglyceride levels at all time points studied. After 14 days on the MCD diet, TTR-Abcb11 mice have 3-fold increases in TNF-alpha mRNA and 3.9-fold increases in IL-6 mRNA compared with FVB/NJ mice. TTR-Abcb11 mice also had a greater increase in cytochrome P-450 2E1 expression. A greater decrease in sterol regulatory element binding protein-1c and fatty acid synthase mRNA expression was also seen in TTR-Abcb11 compared with wild-type mice fed an MCD diet. They also have enhanced TNF-alpha, IL-6, and cytochrome P-450 2E1 expression. We conclude that TTR-Abcb11 mice develop a more rapid hepatitis with less steatosis.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2005

Hemodynamics of distal revascularization-interval ligation.

Distal revascularization-interval ligation (DRIL) empirically corrects steal after arteriovenous fistula (AVF) creation in most cases, but because there is no topologic alteration in anatomy, it is unclear as to why it is effective. To explore this issue, nine symptomatic patients underwent intravascular pressure and flow measurements before and after DRIL following upper arm autologous AVFs. Mean pre-DRIL systolic pressure (mmHg; mean +/- SD) in the proximal brachial artery (PROX) was 102 +/- 17, while that at the AV anastomosis (AV ANAST) was 47 +/- 38 (p < 0.0006). Flow (mL/min) distal to AV ANAST was retrograde with the fistula open (-21 +/- 64) but became antegrade (58 +/- 29; p < 0.03) with occlusion of the fistula. Following DRIL, pressures at both PROX and AV ANAST sites did not change (104 +/- 24 and 51 +/- 43, respectively). However, pressure at the point at which the blood flow split to supply the hand or the fistula, now PROX, increased from 47 +/- 38 (pre-DRIL AV ANAST) to 104 +/- 24 (p < 0.0001). Pressure in the brachial artery distal to the ligature increased to 104 +/- 27 (p < 0.0001), flow at this point (to the hand) became antegrade (51 +/- 39; p < 0.03), and occlusion of the fistula did not significantly change pressure at this site. We hypothesize that improvement in hand perfusion following DRIL is due to a higher pressure at the point at which the blood flow splits to supply both hand and fistula (pre-DRIL: AV ANAST; post-DRIL: PROX), allowing antegrade flow down the new bypass to the lower pressure forearm. This increased pressure must be due to the increased resistance of the fistula created by interposing the arterial segment between the original AV ANAST and new PROX ANAST. As such, DRIL is schematically equivalent to banding, but resistance is increased in a fashion that is physiologically and empirically acceptable.

Aged↗

PubMed2005

Percutaneous and open renal revascularizations have equivalent long-term functional outcomes.

Atherosclerotic renal artery stenosis is a significant cause of poorly controlled hypertension and progressive renal dysfunction leading to ischemic nephropathy and other end-organ damage. The optimal treatment of renovascular disease contributing to hypertension and renal dysfunction is not known. This study compares the anatomic and functional outcomes of both open and endovascular therapy for chronic, symptomatic atherosclerotic renal artery disease. We performed a retrospective analysis of records from patients who underwent renal arterial interventions, endovascular or open bypass, between January 1984 and January 2004. Principal indications for intervention were hypertension (51%), chronic renal insufficiency (13%), and hypertension and elevated creatinine (36%). A total of 247 patients (109 males; mean age 69 +/- 10, range 44-89 years) underwent 314 interventions (109 open procedures; 205 angioplasties, 71% with stent placement). There was a significant difference in 30-day mortality (4% vs. <1%; p < 0.005) between the open and endoluminal groups, but not at 1, 3, or 5 years. Patients in the open group had a higher primary patency rate at 5 years (83 +/- 5% vs. 76 +/- 6%; p = 0.03), but patients in the endoluminal group had a higher assisted primary patency rate at 5 years (92 +/- 5% vs. 84 +/- 5; p = 0.03). There was no significant difference between both treatment groups in cumulative freedom from presenting symptom or in freedom from dialysis and renal-related death. Patients who presented with hypertension were more likely to have shown improvement in their blood pressure with endoluminal intervention at 1, 3, and 5 (59 +/- 6% endoluminal vs. 83 +/- 5% open; p = 0.01) years. From these results we conclude that open repair and endoluminal repair of atherosclerotic renal artery stenosis have similar immediate and long-term functional and anatomic outcomes. Patients who present with hypertension may have greater benefit with an endoluminal repair.

Aged↗

PubMed2005

Percutaneous angioplasty and stenting of the superficial femoral artery.

OBJECTIVES: The objectives of this study were to examine factors predictive of success or failure after percutaneous angioplasty (PTA) and stenting (S) of the superficial femoral artery (SFA) and to compare the results of PTA/S with a contemporary group of patients treated with femoropopliteal bypass. METHODS: A database of patients undergoing PTA and/or S of the SFA between 1986 and 2004 was maintained. Intention-to-treat analysis was performed. Patients underwent duplex scanning follow-up at 1, 3, and every 6 months after the intervention. Angiograms were reviewed in all cases to assess lesion characteristics and preprocedure and postprocedure runoff. Results were standardized to current TransAtlantic Inter-Society Consensus (TASC) and Society for Vascular Surgery (SVS) criteria. Kaplan-Meier survival analyses were performed to assess time-dependent outcomes. Cox proportional hazard analyses were performed to assess factors associated with patient survival and treatment efficacy. RESULTS: Three hundred eighty total limbs underwent PTA/S in 329 patients (67% male, 33% female; average age, 65 years). Mean follow-up was 1.8 years from the date of initial intervention. Indications for intervention were claudication in 66%, rest pain in 16%, and tissue loss in 18%. Runoff at the tibial level was 2.1 +/- 0.8 patent vessels. Mean SVS ischemia grade was 3.1 (range, 1 to 5). TASC lesion grades were A (48%), B (18%), C (22%), and D (12%). Angioplasty alone was used in 63% of cases. Primary treatment failure (inability to cross lesion) was seen in 7% of patients. There was one periprocedural death. Primary patency rates were 86% at 3 months, 80% at 6 months, 75% at 12 months, 66% at 24 months, 60% at 36 months, 58% at 48 months, and 52% at 60 months. Assisted primary patency rates were slightly higher ( P = not significant). By Cox proportional hazards analysis, patency of PTA/S was associated with higher preoperative ankle/brachial index ( P = .016) and the performance of angioplasty only ( P = .011). Failed or occluded PTA/S was associated with TASC C ( P < .0001) and TASC D lesions ( P < .0001). Patient death was associated with the presence of congestive heart failure ( P = .003). Subgroup analysis revealed that primary patency rates are highly dependent on lesion type (A > B > C > D, P < .0001). PTA/S patency for TASC A and B lesions compared favorably to prosthetic and venous femoropopliteal bypass. Surgical bypass was superior to PTA/S for TASC C and D lesions. CONCLUSIONS: PTA and stenting of the SFA can be performed safely with excellent procedural success rates. Improved patency of these interventions was seen with increased ankle/brachial index and the performance of angioplasty only. Worse patency was seen with TASC C and TASC D lesions. Patency rates were strongly dependent on lesion type, and the results of angioplasty and stenting compared favorably with surgical bypass for TASC A and B lesions.

Aged↗

PubMed2005

Management of complex groin wounds: preferred use of the rectus femoris muscle flap.

This study reviews our experience with the rectus femoris muscle flap for complex groin wound reconstruction. Over the past 5 years, the rectus femoris has become our routine method of groin wound reconstruction. The rectus femoris is harvested through a midanterior incision extending over the distal two-thirds of the thigh. The muscle is elevated on its pedicle and transposed into the groin wound defect either directly or through an intervening skin bridge. Hospital and outpatient records were reviewed for all patients undergoing groin wound reconstruction with this technique from 1999 through 2003. Thirty-seven rectus femoris muscle flaps were performed in 33 patients. The mean patient age was 65.3 years (range, 25 to 88 years). Thirty groin wounds (81.1 percent) occurred after infrainguinal revascularization, 23 (76.7 percent) of which contained prosthetic material. Five (21.7 percent) of these wounds had their prosthetic material removed at the time of reconstruction. The remaining seven groin wounds (18.9 percent) occurred after femoral vessel cannulation for either cardiac or transplant surgery. There were no intraoperative mortalities and no anastomotic hemorrhages. There were no flap losses. Thirty-five of the 37 treated wounds healed (94.6 percent), 26 primarily (70.3 percent) and nine (24.3 percent) after delayed healing and contracture. Reoperation was performed in one patient for flap readvancement and in three patients for prosthetic graft removal after initial flap reconstruction. Two patients (6.1 percent) died during their hospitalization with persistent open groin wounds after flap reconstruction. All muscle flap donor incisions healed, with only two (5.4 percent) experiencing minimal delayed healing. There were no donor-site wound infections and no donor sites required reoperation. Thirty-three groin wounds (89.2 percent) demonstrated culture-positive microbial infection, 15 (45.5 percent) of which were polymicrobial. The 30-day mortality rate was 15.2 percent and the 6-month mortality rate increased to 27.2 percent, with multisystem organ failure as the most common cause. The rectus femoris muscle flap is an effective and reliable means of complex groin wound reconstruction. The muscle flap is dependable and the donor site is not problematic, even in the presence of peripheral vascular disease. On the basis of our clinical results, we believe that the rectus femoris muscle flap is the flap of choice for groin wound reconstruction.

Adult↗

PubMed2005

Stacked proximal aortic cuffs: an "off-the-shelf" solution for treating focal thoracic aortic pathology.

PURPOSE: To report our early experience with the endovascular placement of stacked Zenith main body extensions (cuffs) in the treatment of focal thoracic aortic pathology in high-risk patients. METHODS: Between January 2003 and May 2004, 6 patients (3 men; mean age 59 years, range 37-82) with focal aortic pathology underwent endovascular repair using stacked 30 and 32-mm-diameter Zenith main body extensions. The setting was a university tertiary referral center for vascular disease. Indication for treatment included 2 descending thoracic aneurysms and individual cases of traumatic thoracic tear, diverticulum of Kommerell, thoracic pseudoaneurysm, and aortoesophageal fistula. RESULTS: All procedures were performed successfully, with a mean of 3 cuffs used. The patient with an aortoesophageal fistula expired after successful cuff placement due to sequela of massive pretreatment hemorrhage; fistula coverage was confirmed at autopsy. There were no type I endoleaks. Morbidity included an occluded right subclavian artery from traumatic passage of the device through the artery. No left subclavian arteries were covered. No neurological deficits or paraplegia was observed. The cuffs were patent in all surviving patients at an average follow-up of 7 months (range 3-12). Computed tomography in all survivors confirmed adequate cuff placement, absence of endoleak, and lack of cuff migration. Based on this experience, the following technical recommendations are offered: (1) right subclavian cutdown when needed to reach a lesion beyond the range of the sheath, (2) Dacron chimney placement, (3) stiff guidewire usage, (4) wire placement from the right subclavian artery through the common femoral artery if necessary to ease a sharp bend in the arch, and (5) cuff overlap of 25% to 50%. CONCLUSIONS: In high-risk patients, focal aortic pathology can be successfully treated with off-the-shelf commercially available cuffs using a stacking technique with acceptable mortality, morbidity, and short-term durability.

Adult↗

PubMed2004

Transcriptional regulation of murine Slc22a1 (Oct1) by peroxisome proliferator agonist receptor-alpha and -gamma.

The transport and metabolism of organic cationic endobiotics, nutrients, and drugs are essential hepatic functions. Slc22A1 is the basolateral liver transporter mediating the uptake of organic cations; however, little is known about the regulation of this transport protein. Peroxisome proliferator agonist receptor (PPAR)-alpha and -gamma agonists are commonly used agents that regulate many hepatocellular transport functions. Thus the purpose of this study was to examine the effects of PPAR agonists on the hepatic regulation and function of Slc22a1. Mice and H35 cells were administered PPAR-alpha and -gamma agonists, and the effect on Slc22a1 gene expression was measured. We subsequently cloned the Slc22a1 promoter and employed chimeric constructs to assay Slc22a1 gene transcription. The effects of PPAR-agonist treatment on organic cation uptake was also assayed. Slc22a1 expression was increased by PPAR-alpha and -gamma agonist treatment in both murine livers and H35 cells. Gene expression in H35 cells was further increased following transfection with expression vectors of PPAR transcription factors and PPAR agonist treatment. We cloned the promoter region of Slc22a1 and identified a PPAR-response element, and transfections with chimeric Slc22a1; promoter-reporter gene constructs demonstrate that the increased gene expression was transcriptionally regulated. Functional assays confirmed that cells treated with PPAR agonists displayed significant increases in organic cation uptake. PPAR-alpha and -gamma agonists transcriptionally increase Slc22a1 gene expression, and the increased Slc22a1 expression results in enhanced cellular organic cation uptake. These studies may have implication for the uptake of organic cationic drugs and for lipid metabolism.

Animals↗

PubMed2004

Obese and diabetic db/db mice develop marked liver fibrosis in a model of nonalcoholic steatohepatitis: role of short-form leptin receptors and osteopontin.

Obesity and type 2 diabetes are associated with nonalcoholic steatohepatitis (NASH), but an obese/diabetic animal model that mimics human NASH remains undefined. We examined the induction of steatohepatitis and liver fibrosis in obese and type 2 diabetic db/db mice in a nutritional model of NASH and determined the relationship of the expressions of osteopontin (OPN) and leptin receptors to the pathogenesis of NASH. db/db mice and the corresponding lean and nondiabetic db/m mice were fed a diet deficient in methionine and choline (MCD diet) or control diet for 4 wk. Leptin-deficient obese and diabetic ob/ob mice fed similar diets were used for comparison. MCD diet-fed db/db mice exhibited significantly greater histological inflammation and higher serum alanine aminotransferase levels than db/m and ob/ob mice. Trichrome staining showed marked pericellular fibrosis in MCD diet-fed db/db mice but no significant fibrosis in db/m or ob/ob mice. Collagen I mRNA expression was increased 10-fold in db/db mice, 4-fold in db/m mice, and was unchanged in ob/ob mice. mRNA expressions of OPN, TNF-alpha, TGF-beta, and short-form leptin receptors (Ob-Ra) were significantly increased in db/db mice compared with db/m or ob/ob mice. Parallel increases in OPN and Ob-Ra protein levels were observed in db/db mice. Cultured hepatocytes expressed only Ob-Ra, and leptin stimulated OPN mRNA and protein expression in these cells. In conclusion, our results demonstrate the development of an obese/diabetic experimental model for NASH in db/db mice and suggest an important role for Ob-Ra and OPN in the pathogenesis of NASH.

Animals↗

PubMed2004

Upregulation of osteopontin expression is involved in the development of nonalcoholic steatohepatitis in a dietary murine model.

The pathogenesis of nonalcoholic steatohepatitis (NASH) is poorly defined. Feeding mice a diet deficient in methionine and choline (MCD diet) induces experimental NASH. Osteopontin (OPN) is a Th1 cytokine that plays an important role in several fibroinflammatory diseases. We examined the role of OPN in the development of experimental NASH. A/J mice were fed MCD or control diet for up to 12 wk, and serum alanine aminotransferase (ALT), liver histology, oxidative stress, and the expressions of OPN, TNF-alpha, and collagen I were assessed at various time points. MCD diet-fed mice developed hepatic steatosis starting after 1 wk and inflammation by 2 wk; serum ALT increased from day 3. Hepatic collagen I mRNA expression increased during 1-4 wk, and fibrosis appeared at 8 wk. OPN protein expression was markedly increased on day 1 of MCD diet and persisted up to 8 wk, whereas OPN mRNA expression was increased at week 4. TNF-alpha expression was increased from day 3 to 2 wk, and evidence of oxidative stress did not appear until 8 wk. Increased expression of OPN was predominantly localized in hepatocytes. Hepatocytes in culture also produced OPN, which was stimulated by transforming growth factor-beta and TNF-alpha. Moreover, MCD diet-induced increases in serum ALT levels, hepatic inflammation, and fibrosis were markedly reduced in OPN(-/-) mice when compared with OPN(+/+) mice. In conclusion, our results demonstrate an upregulation of OPN expression early in the development of steatohepatitis and suggest an important role for OPN in signaling the onset of liver injury and fibrosis in experimental NASH.

Alanine Transaminase↗

PubMed2004

Rotational muscle flap closure for acute groin wound infections following vascular surgery.

Since 1996, 41 patients have presented to our institution with deep but localized groin infection following bypass (30) or isolated femoral artery surgery (11). These patients were treated with antibiotics, debridement, and rotational muscle flap coverage either immediately or within a few days. Patients had one of three patterns: serous leak from a groin incision within a few days of operation (Acute, n = 10), early serous leak that later became grossly infected (Acute-observed, n = 8), or obvious purulent drainage following an initially normal, healed wound (Delayed, n = 23). Patients with early leak had nearly uniformly polymicrobial infections with a preponderance of gram-negative organisms, whereas most of those with late purulence had monobacterial infection with Staphylococcus aureus. At exploration, 26 of 41 suture lines were exposed. Rectus femoris flaps were used in 35 patients (85% of cases) for coverage, and graft preservation was attempted in all 8 vein grafts and 16 of 23 prosthetic grafts. Only one flap failed and there were no instances of anastomotic bleeding. There were no deaths directly attributable to reexploration and flap coverage, although 10 patients died during the index hospitalization. Durable coverage with no long-term evidence of infection was achieved in 24 patients with mean follow-up of 23 (range 10-66) months and another 12 had no evidence of local problems despite shorter follow-up; only 5 patients (12%) overall had evidence of persistent graft infection or unexplained bacteremia. In patients with attempted graft salvage, limb salvage was 97% at 6 months and 85% at 1 year. Although early mortality is high, deaths are not related to the flap procedure itself, local outcome is excellent, and graft and limb salvage are good; results are much worse if an initially draining wound is treated too late. Local rotational muscle flap closure is an excellent solution for acute infections involving the groin following vascular procedures.

Blood Vessel Prosthesis↗

PubMed2004

The methionine-choline deficient dietary model of steatohepatitis does not exhibit insulin resistance.

BACKGROUND/AIMS: Non-alcoholic steatohepatitis is an important disease whose pathophysiology remains incompletely understood, although in humans a strong association with insulin resistance exists. Mice fed a methionine-choline deficient (MCD) diet develop steatohepatitis, however the influence of insulin in this model is unknown. METHODS: Male FVB/NJ mice were fed the MCD, MCD control or chow diet for 10 or 28 days. Fasting glucose, ALT, triglyceride and insulin was measured. Glucose tolerance tests (GTT) and insulin tolerance tests (ITT) were performed followed by quantitative insulin sensitivity check index (QUICKI) determination. RESULTS: ALT levels were significantly higher in the MCD group. Fasting glucose was 81+/-5 mg/dl in MCD diet fed mice, compared to MCD controls (196+/-46 mg/dl) and chow (199+/-15 mg/dl) (P<0.0001). During GTT and ITT, the effect of glucose administration on blood glucose was dampened, and the insulin effect more pronounced in the MCD group (P=0.026 and P<0.001). QUICKI in MCD fed mice was significantly higher than in the chow fed mice. CONCLUSIONS: GTT, ITT and QUICKI confirmed the absence of insulin resistance in the MCD fed mice. This model causes fibrosing steatohepatitis and may help delineate the non-insulin resistant mechanisms involved in human steatohepatitis.

Alanine Transaminase↗

PubMed2004

Pentoxifylline attenuates steatohepatitis induced by the methionine choline deficient diet.

BACKGROUND/AIMS: Feeding mice a methionine choline deficient (MCD) diet serves as a nutritional model of non-alcoholic steatohepatitis (NASH). NASH and alcohol-induced steatohepatitis are histologically similar, suggesting a similar pathogenesis. Pentoxifylline (PTX) attenuates TNF-alpha production, acts as an antioxidant and decreases mortality in alcoholic steatohepatitis. The aim of our study is to determine if PTX attenuates MCD diet induced steatohepatitis and determine the mechanism of this effect. METHODS: Mice were placed on an MCD or control diet for 2 weeks and were treated with or without PTX. Serum ALT, liver histology, and inflammatory mechanisms were evaluated. RESULTS: PTX attenuates MCD diet induced steatohepatitis, decreasing both serum ALT levels and hepatic inflammation. Serum ALT levels were reduced approximately 50% in the MCD+PTX group compared to the MCD group. Hepatic glutathione levels were significantly higher in the MCD+PTX group compared to the MCD group. There was also a reduction in TNF-alpha mRNA in female mice treated with PTX. MCD+PTX mice had increased hepatic triglyceride content compared to the MCD mice, but less histologic evidence of inflammation despite the increased steatosis. Serum lipid and bile salt levels also were similar in PTX and vehicle control treated mice. CONCLUSIONS: PTX decreases serum ALT levels and hepatic inflammation in the MCD model of steatohepatitis, likely via increasing glutathione levels or reducing TNF-alpha expression.

Alanine Transaminase↗