Relationship between lipogenesis,ketogenesis, and malonyl-CoA content in isolated hepatocytes from the obese Zucker rat adapted to a high-fat diet

The relationship between lipogenesis and ketogenesis and the concentration of malonyl coenzyme A (CoA) was investigated in hepatocytes from adult obese Zucker rats and their lean littermates fed either a control low-fat diet or a high-fat diet (30% lard in weight). With the control diet, lipogenesis—although strongly inhibited in the presence of either 1 mmol/L oleate, 10^?6 mol/L glucagon or 0.1 mmol/L TOFA (a hypolipidemic drug)—remained about fifteen-fold higher in the obese rats than in the lean rats. In contrast, ketogenesis under some conditions (oleate + TOFA) was not significantly lower (30%) as compared with the lean rats. After adaptation to the high-fat diet, lipogenesis was depressed fourfold in the lean rats and ninefold in the obese ones; however its magnitude remained significantly higher in the latter, namely at a value close to that measured in control-fed lean rats. Ketogenesis was comparable in lean and obese rats and much higher in the presence of 1 mmol/L oleate than of 0.3 mmol/L oleate, whereas lipogenesis did not vary with increasing oleate concentration in the medium. Acetyl-CoA carboxylase activity measured in liver homogenates was higher in the obese group, but was stepwise inhibited by increasing concentrations of oleyl-CoA regardless of the diet for both lean and obese rats, thus showing no abnormality of in vitro responsiveness to this inhibitor. With the control diet, hepatocyte malonyl-CoA levels were significantly higher in the obese rats, both in the basal state and after inhibition of lipogenesis by oleate and TOFA. However, after the high-fat diet, there was no longer a significant difference between the genotypes. These results show that in the obese Zucker rats, ketogenesis is dependent on hepatocyte malonyl-CoA content in the sense that their ketogenic capacity becomes “normalized” when malonyl-CoA is decreased to the levels found in the lean littermates, as it is the case after fat-feeding. This normalization of malonyl-CoA levels in spite of higher lipogenesis in the obese rats may result from the activities of enzymes of its formation and utilization.     

Dynamic MRI after surgical repair for pelvic organ prolapse

PURPOSE: The purpose of this work was to compare dynamic MRI measurement results from patients with pelvic organ prolapse before and after surgical repair and to correlate postoperative MRI findings with the results of physical examination and patients' complaints. MATERIALS AND METHODS: Thirty women were included in this prospective study. Fifteen patients with uterovaginal or vaginal vault descent and 15 asymptomatic female volunteers underwent dynamic MRI using a single-shot, fast spin-echo sequence with half-Fourier data acquisition. Single-slice midsagittal images of pelvic organs were obtained at maximal pelvic strain and at perineal contraction. Dynamic MRI was repeated on all patients 2-4 months after surgical repair. Postoperative and preoperative dynamic MRIs were compared with the normal values of the volunteer group. Postoperative MRIs were correlated with the clinical findings after surgical repair. RESULTS: Seven patients reported symptoms after surgical repair, although only two had pathologic findings on physical examination (residual urine associated with third-degree stress urinary incontinence, and bladder neck descent). MRI showed pathologic changes in five of these patients. Two patients with persistent first-degree stress incontinence had no pathologic findings on MRI. CONCLUSION: Dynamic MRI is helpful in the evaluation of persistent patient complaints after surgery for pelvic organ prolapse and detects more defects than physical examination does.     

Prediction of necrosis after chemotherapy of advanced germ cell tumors: results of a prospective multicenter trial of the German Testicular Cancer Study Group

PURPOSE: We evaluated the prognostic parameters of necrotic residual tumors after chemotherapy of advanced germ cell tumors to improve on the current indications for surgery. MATERIALS AND METHODS: Between January 1996 and January 2000, in 8 centers of the German Testicular Cancer Study Group, preoperative parameters were assessed to predict necrosis in the residual tumors of 261 patients with retroperitoneal residual tumor resection after first (92%) and second line (8%) chemotherapy. RESULTS: Of 232 evaluable patients 39 had pure seminoma and 5 had viable cancer (1 with seminoma) in the residual tumor. Of the remaining 193 patients with nonseminoma 35% had necrosis, 34% teratoma and 31% had viable carcinoma in the residual tumor. After multivariate analysis and exclusion of patients with seminoma, the 3 parameters independently predictive of necrosis were alpha-fetoprotein before chemotherapy less than 20 ng/ml, and tumor volume before and after chemotherapy. A mathematical model to predict necrosis yielded a test accuracy of 75%, a sensitivity to predict necrosis of 52% and a specificity of 87%. CONCLUSIONS: Patients with pure seminoma should not undergo residual tumor resection because 97% of patients who received adequate chemotherapy were found to have no residual seminoma. In cases of nonseminoma alpha-fetoprotein values before chemotherapy less than 20 ng/ml and a high percentage of shrinkage during chemotherapy reliably predicted only 19% of cases of necrosis. Therefore, this model is clinically irrelevant and patients with minimal residual disease should undergo surgery. New methods are necessary to improve the preoperative selection of patients after chemotherapy.     

Mycophenolate mofetil and sirolimus as calcineurin inhibitor-free immunosuppression for late cardiac-transplant recipients with chronic renal failure

BACKGROUND: Calcineurin-inhibitor (CNI)-related renal failure is a common problem after cardiac transplantation (HTx). The aim of this study was to introduce a CNI-free immunosuppressive regimen to HTx recipients with late posttransplant renal impairment and to evaluate the impact of conversion to this new immunosuppression (mycophenolate mofetil [MMF] and sirolimus [Sir]) treatment on renal function. METHODS AND RESULTS: Thirty-one HTx patients (25 men, 6 women; 0.2-14.2 years after transplantation) with CNI-based immunosuppression and a serum creatinine greater than 1.9 mg/dL were included in the study. Creatinine and cystatin levels were monitored to detect renal function. Mean patient age was 50+/-14 (range 19-74) years. Conversion was started with 6 mg Sir, continued with 2 mg, and the dose was adjusted to achieve target trough levels between 8 and 14 ng/mL. MMF was continued with trough level adjusted (1.5-4 microg/mL). Subsequently, the CNIs were tapered down and stopped. Clinical follow-up (first and every 3 months after conversion) included endomyocardial biopsies, echocardiography, and laboratory studies. Survival was 90% after a mean follow-up of 13+/-95 months. No acute rejection episode was detected during the study period. Renal function improved significantly after conversion: creatinine preconversion vs. postconversion: 3.14+/-0.76 mg/dL vs. 2.14+/-0.83 mg/dL, P =0.001. Cystatin preconversion vs. postconversion: 2.95+/-1.06 mg/L vs. 2.02+/-1.1 mg/L, P =0.01. In three patients, hemodialysis therapy was stopped completely after conversion. Graft function remained stable. Fractional shortening preconversion vs. postconversion: 36.9+/-6% vs. 36.4+/-6%. There were no serious adverse events. One patient had to be excluded because of noncompliance. CONCLUSIONS: Conversion from CNI-based immunosuppression to MMF and Sir in HTx patients with chronic renal failure was safe, preserved graft function, and improved renal function.     

VEGF Improves Skeletal Muscle Regeneration After Acute Trauma and Reconstruction of the Limb in a Rabbit Model

Background

Complicated tibial fractures with severe soft tissue trauma are challenging to treat. Frequently associated acute compartment syndrome can result in scarring of muscles with impaired function. Several studies have shown a relationship between angiogenesis and more effective muscle regeneration. Vascular endothelial growth factor (VEGF) is associated with angiogenesis but it is not clear whether it would restore muscle force, reduce scarring, and aid in muscle regeneration after acute musculoskeletal trauma.

Questions/purposes

Therefore, we asked whether local application of VEGF (1) restores muscle force, (2) reduces scar tissue formation, and (3) regenerates muscle tissue.

Methods

We generated acute soft tissue trauma with increased compartment pressure in 22 rabbits and shortened the limbs to simulate fracture débridement. In the test group (n = 11), a VEGF-coated collagen matrix was applied locally around the osteotomy site. After 10 days of limb shortening, gradual distraction of 0.5 mm per 12 hours was performed to restore the original length. Muscle force was measured before trauma and on every fifth day after trauma. Forty days after shortening we euthanized the animals and histologically determined the percentage of connective and muscle tissue.

Results

Recovery of preinjury muscle strength was greater in the VEGF group (2.4 N; 73%) when compared with the control (1.8 N; 53%) with less connective and more muscle tissue in the VEGF group. The recovery of force was related to the percentage of connective tissue versus muscle fibers.

Conclusions

Local application of VEGF may improve restoration of muscle force by reducing connective tissue and increasing the relative amount of muscle fibers.

Clinical Relevance

VEGF may be useful to improve skeletal muscle repair by modulating muscle tissue regeneration and fibrosis reduction after acute trauma.     

Surgical epicardial left ventricular lead versus coronary sinus lead placement in biventricular pacing.

OBJECTIVE: Biventricular pacing has demonstrated improvement in cardiac function in treating congestive heart failure (CHF). Two different operative strategies (coronary sinus vs. epicardial stimulation) for left ventricular (LV) pacing were compared. METHODS: Since April 1999, a total of 86 patients (pts, age: 63+/-10 years) with depressed systolic LV function (mean ejection fraction 24+/-9%), left bundle-branch-block (mean QRS 182+/-22 ms) and congestive heart failure NYHA III or higher were enrolled. For biventricular stimulation coronary sinus (CS) leads were placed in 79 pts. Nine of these devices were converted to surgical epicardial LV-leads, because of CS-lead failure. In 7 patients epicardial LV-leads were initially implanted surgically, accounting for a total of 16 pts with surgical placed epicardial steroid-eluting LV-leads. For these, a limited left-lateral thoracotomy (7+/-4 cm) was used. Thirty-three (38%) pts had an indication for a defibrillator. The mean follow-up time was 16.4+/-15.4 months (0.1-45 months), representing 107.1 patient-years. RESULTS: In the biventricular pacing mode, QRS duration decreased to 143+/-16 ms (P<0.001). Threshold capture of the CS-leads increased significantly compared to surgically placed epicardial leads (18 month control: 2.2+/-1.4V/0.5 ms vs. 0.7+/-0.3V/0.5 ms), which had no increase in threshold (P<0.001). At the 18 month follow-up 7 CS-leads had a threshold of >4V/0.5 ms vs. epicardial leads which were under 1.1V/0.5 ms, except for one (1.8V/0.5 ms). After CS-lead implantation 25 LV-lead related complications occurred, (failed implantation, CS-dissection, loss of pacing capture, diaphragm stimulation or lead dislodgment), vs. one dislodgement after surgical epicardial lead placement (P<0.05). Correct lead positioning (obtuse marginal branch area) was achieved in all surgical epicardial placements but only in 70% with CS-leads (P<0.03). In the follow up period, 9 pts died (4 cardiac related). Heart transplantation was necessary in 4 pts due to deterioration of the cardiomyopathy. CONCLUSIONS: Surgical epicardial lead placement revealed excellent long-term results and a lower LV-related complication rate compared to CS-leads. Although, the approach via limited thoracotomy for biventricular pacing is associated with 'more surgery', it is a safe and reliable technique and should be considered as an equal alternative.     

Neovascularization in prefabricated flaps using a tissue expander and an implanted arteriovenous pedicle

Creating prefabricated flaps using tissue expanders in combination with the implantation of maximal blood flow vascular pedicles into suitable tissue areas represents a new tendency in the reconstruction of large skin defects. In 42 Chinchilla Bastard female rabbits weighing 3,700-4,600 g, skeletonized arteriovenous pedicles with maximal blood flow, dissected from the femoral and saphena magna bundles, were implanted underneath abdominal fasciocutaneous flaps. Oval tissue expanders of 250 ml were placed and fixed on the abdominal wall to expand these prefabricated flaps. The evaluation parameters were macroscopic observation, blood analysis, selective microangiography, histology, and scintigraphy. The study results showed that neovascularization in expanded prefabricated flaps was established from newly formed vessels generated from the implanted pedicles and their vascular connections with the originally available vasculature in the flap. After 20 days of prefabrication, the entirety of the expanded prefabricated flaps was perfused by blood flow supplied from newly implanted arteriovenous pedicles. The study indicated that an expanded prefabricated flap can be successfully created by the simultaneous implantation of a maximal blood flow pedicle in combination with flap expansion.     

Rosiglitazone improves glomerular hyperfiltration, renal endothelial dysfunction, and microalbuminuria of incipient diabetic nephropathy in patients

Microalbuminuria, an early feature of diabetic nephropathy, indicates intrarenal endothelial damage. In type 2 diabetes, microalbuminuria is strongly related to insulin resistance. We therefore investigated whether rosiglitazone, an insulin-sensitizing drug that is known to improve endothelial dysfunction, was able to improve intrarenal endothelial dysfunction and microalbuminuria. Nineteen type 2 diabetic patients participated in this double-blind cross-over trial. Nine patients with newly diagnosed disease without microalbuminuria were randomized to a treatment with rosiglitazone or nateglinide, each for 12 weeks. Ten patients with microalbuminuria were randomized to rosiglitazone or placebo, each for 12 weeks in addition to their previous antidiabetic medication. After each treatment, glomerular filtration rate (GFR), renal plasma flow, and filtration fraction were measured before and after blockade of nitric oxide (NO) by intravenous administration of N-monomethyl-L-arginine-acetate (L-NMMA). Ten healthy subjects served as control subjects. Type 2 diabetic patients at baseline showed glomerular hyperfiltration compared with healthy control subjects. Rosiglitazone reduced elevated GFR and filtration fraction toward control primarily in patients with microalbuminuria (GFR: 133.4 +/- 9.8 vs. 119.6 +/- 8.7 ml/min; filtration fraction: 23.2 +/- 1.7 vs. 20.5 +/- 1.6% before and after rosiglitazone, respectively; control subjects: GFR 111.7 +/- 8.6 ml/min, filtration fraction 20.4 +/- 1.5%). Rosiglitazone improved intrarenal NO bioavailability in type 2 diabetes toward control as shown by infusion of L-NMMA. Rosiglitazone reduced albumin excretion in type 2 diabetes with microalbuminuria from 116.5 +/- 31 to 40.4 +/- 12 mg/day. Rosiglitazone ameliorated glomerular hyperfiltration in early type 2 diabetes, improved NO bioavailability, and lessened renal end-organ damage in type 2 diabetes with microalbuminuria.     

Prospective cohort study of effects of infliximab on rheumatoid factor,anti-cyclic citrullinated peptide antibodies and antinuclear antibodies in patients with long-standing rheumatoid arthritis

BackgroundAntibodies to cyclic citrullinated peptide (anti-CCP) and IgM rheumatoid factor (IgM-RF) are well-established serological markers for rheumatoid arthritis (RA). Lupus-like disease with antinuclear antibodies (ANA) has been reported during TNFα antagonist therapy. Our objectives were to investigate the effect of infliximab therapy on these three autoantibodies in patients with established RA and to look for correlations linking IgM-RF and anti-CCP titres to a treatment response (defined as a good or moderate EULAR response) after 48 weeks of infliximab therapy.MethodsThirty-six patients with long-standing RA not responding to disease-modifying anti-rheumatic drugs (DMARDs) received intravenous infliximab (starting dose: 3 mg/kg) at 0, 2, and 6 weeks then at 8-week intervals, in combination with a DMARD. At baseline, week 24, and week 48, C-reactive protein (CRP) and the erythrocyte sedimentation rate (ESR) were determined and the disease activity score (DAS28) was calculated. Serum samples collected at the same time points were used to measure anti-CCP (commercial second-generation ELISA), IgM-RF (quantitative nephelometric assay), and ANA (indirect immunofluorescence in HEp2 cells). Correlations linking baseline autoantibody titres to changes in autoantibody levels were examined.ResultsAt baseline, tests were positive for anti-CCP in 31/36 (94.6%) patients, IgM-RF in 29/36 (80.5%) patients, and ANA in 16/36 (44%) patients. IgM-RF titres decreased significantly (p < 0.001), whereas anti-CCP showed little change (p = 0.053). ANA titres increased significantly (p < 0.001). The treatment response was not associated with changes in anti-CCP or IgM-RF titres during infliximab therapy (OR for a response in patients with a 50% anti-CCP decrease, 0.77 [95%CI, 0.16–3.58]; OR for a response in patients with a 50% IgM-RF decrease, 0.82 [95%CI, 0.16–4.13]).ConclusionsDuring infliximab therapy used to treat established RA, IgM-RF titres showed larger decreases than anti-CCP titres. Changes in IgM-RF and anti-CCP failed to correlate with the 48-week treatment response.     

Molecular and functional expression of voltage-operated calcium channels during osteogenic differentiation of human mesenchymal stem cells.

We used the patch-clamp technique and RT-PCR to study the molecular and functional expression of VOCCs in undifferentiated hMSCs and in cells undergoing osteogenic differentiation. L-type Ca2+ channel blocker nifedipine did not influence alkaline phosphatase activity, calcium, and phosphate accumulation of hMSCs during osteogenic differentiation. This study suggests that osteogenic differentiation of hMSCs does not require L-type Ca2+ channel function. INTRODUCTION: During osteogenic differentiation, mesenchymal stem cells from human bone marrow (hMSCs) must adopt the calcium handling of terminally differentiated osteoblasts. There is evidence that voltage-operated calcium channels (VOCCs), including L-type calcium channels, are involved in regulation of osteoblast function. We therefore studied whether VOCCs play a critical role during osteogenic differentiation of hMSCs. MATERIALS AND METHODS: Osteogenic differentiation was induced in hMSCs cultured in maintenance medium (MM) by addition of ascorbate, beta-glycerophosphate, and dexamethasone (ODM) and was assessed by measuring alkaline phosphatase activity, expression of osteopontin, osteoprotegerin, RANKL, and mineralization. Expression of Ca2+ channel alpha1 subunits was shown by semiquantitative or single cell RT-PCR. Voltage-activated calcium currents of hMSCs were measured with the whole cell voltage-clamp technique. RESULTS: mRNA for the pore-forming alpha1C and alpha1G subunits of the L-type and T-type Ca2+ channels, respectively, was found in comparable amounts in cells cultured in MM or ODM. The limitation of L-type Ca2+ currents to a subpopulation of hMSCs was confirmed by single cell RT-PCR, where mRNA for the alpha1C subunits was detectable in only 50% of the cells cultured in MM. Dihydropyridine-sensitive L-type Ca2+ currents were found in 13% of cells cultured in MM and in 12% of the cells cultured in ODM. Under MM and ODM culture conditions, the cells positive for L-type Ca2+ currents were significantly larger than cells without Ca2+ currents as deduced from membrane capacitance; thus, current densities were comparable. Addition of the L-type Ca2+ channel blocker nifedipine to the culture media did not influence alkaline phosphatase activity and the extent of mineralization. CONCLUSION: These results suggest that, in the majority of hMSCs, Ca2+ entry through the plasma membrane is mediated by some channels other than VOCCs, and blockade of the L-type Ca2+ channels does not affect early osteogenic differentiation of hMSCs.