Catecholamine-induced oligodendrocyte cell death in culture is developmentally regulated and involves free radical generation and differential activation of caspase-3

Oligodendrocyte cultures were used to study the toxic effects of catecholamines. Our results showed that catecholamine-induced toxicity was dependent on the dose of dopamine or norepinephrine used and on the developmental stage of the cultures, with oligodendrocyte progenitors being more vulnerable. A role for oxidative stress and apoptosis on the mechanism of action of catecholamines on oligodendrocyte cell death was next assessed. Catecholamines caused a reduction in intracellular glutathione levels, an accumulation in reactive oxygen species and in heme oxygenase-1, the 32 kDa stress-induced protein. All these changes were prevented by N-acetyl-L-cysteine, a thiocompound with antioxidant activity and a precursor of glutathione, and were more pronounced in progenitors than mature cells, which could contribute to their higher susceptibility. Apoptotic cell death, as assessed by activation of caspase-9 and -3 and cleavage of poly(ADP-ribose) polymerase (a substrate of caspase-3), was only observed in oligodendrocyte progenitors. Pretreatment with zVAD, a general caspase inhibitor, prevented activation of caspase-9 and -3, DNA fragmentation, and decreased progenitors cell death. Furthermore, the expression levels of procaspase-3 and the ratio of the proapoptotic protein bax to antiapoptotic protein bcl-xl were several folds higher in immature than mature oligodendrocytes. Taken together, these results strongly suggest that the catecholamine-induced cytotoxicity in oligodendrocytes is developmentally regulated, mediated by oxidative stress, and have characteristics of apoptosis in progenitor cells.     

In vitro evaluation of the quality of essential drugs on the Tanzanian market

We evaluated the in vitro availability and its stability under simulated tropical conditions of various formulations of four essential drugs marketed in Tanzania. We obtained 22 formulations (containing paracetamol, acetylsalicylic acid, chloroquine or sulphadoxine/pyrimethamine) from wholesale pharmacies in Dar es Salaam and the Medical Stores Department (Tanzania). The drug content, in vitro availability (dissolution) and its stability under simulated tropical conditions were determined using methods specified in the United States Pharmacopoeia (USP) 24 monograph of the respective drugs. All formulations passed the pharmacopoeia requirements for the drug content. However, seven formulations (three acetylsalicylic acid, two sulphadoxine/pyrimethamine and two paracetamol) failed to meet the USP 24 tolerance limits for dissolution. Another five formulations (three paracetamol and two chloroquine) failed to meet the dissolution tolerance limits after being subjected to an accelerated stability test under simulated tropical conditions (75% RH/40 degrees C) for 6 months. The study has demonstrated the presence on the Tanzanian market of essential drug formulations that met potency requirements and yet had unsatisfactory in vitro availability as they were not robust enough to withstand storage under simulated tropical conditions.     

Nutritional rickets and z scores for height in the United Arab Emirates: To D or not to D?

Background: Vitamin D deficiency is still prevalent worldwide, including the Middle East. A cohort of patients with nutritional rickets was treated with vitamin D₂ (ergocalciferol) alone. After this intervention, patients were followed to document changes in z scores for height after treatment. The secondary aim was to determine the proportion of affected children who had vitamin D deficiency or calcium deficiency.
Methods: Z score for height was calculated as the difference between the observed value and the median value, divided by the SD of the population. Z scores were compared in patients before and after treatment.
Results: The improvement in z score after treatment was 0.86 ± 0.95. The 95% confidence interval for the mean difference was 1.32–0.40 ( t  = 3.95, P  < 0.001). With a diagnostic cut-off for 25 hydroxyvitamin D₃ (25D) deficiency of <25 nmol/L, only half were diagnosed with severe vitamin D deficiency. The remaining patients had presumable calcium deficiency. The alkaline phosphatase (ALP) was negatively correlated to z scores, implying that higher ALP concentrations predicted severe bone disease (lower z scores). The variables 25D and age were moderately and positively correlated (Pearson's r  = 0.59, 95%CI: 0.15–0.84; P  = 0.01), indicating that younger infants had the lowest 25D levels.
Conclusion: Vitamin D alone was efficient in resolving radiological and biochemical disturbances as well as improving z scores for height in a cohort of children with nutritional rickets, which included patients with 25D deficiency as well as calcium deficiency. The results support the hypothesis of the interplay and continuum of 25D deficiency and calcium deficiency in the pathogenesis of rickets.     

The promoter region (-800, -509) polymorphisms of transforming growth factor-beta1 (TGF-beta1) gene and recurrent spontaneous abortion

Recurrent spontaneous abortion (RSA) is regarded as a common pregnancy complication in southern Iran. The exact causes of RSA are not yet known. Transforming growth factor-beta1 (TGF-beta1) is produced by T regulatory lymphocytes (Treg), which play an important role in the physiology of pregnancy. Several polymorphisms of the TGF-beta1 gene have been reported, some with important correlation with disease severity. In this investigation, the polymorphism of the TGF-beta1 gene at promoter region positions -800 (G/A) and -509 (C/T) was studied in 111 RSA and 110 normal female subjects from southern Iran by PCR-RFLP. Results indicated that at position -800 (G/A) polymorphism, 75.7% of RSA cases and 77.3% of normals were homozygote GG. In addition, 23.4% of cases and 22.7% of normal individuals were heterozygote AG. Only one of the patients appeared to be homozygote AA. None of the normal individuals were found to be homozygote AA at this position. In the case of the -509 (C/T) polymorphism, 38.7% of patients and 28.2% of controls were homozygote CC. While 40.6% of cases and 50.9% of normal individuals were heterozygote CT, 20.7% of RSA cases and 20.9% of controls were homozygote TT. The results indicate that there are no statistically significant differences of genotype distribution and allele frequency between RSA cases and controls at both polymorphic sites. In conclusion, the promoter region polymorphisms of TGF-beta1 at positions -800 (G/A) and -509 (C/T) may not be associated with RSA.     

Pro-oxidant activity of zuclopenthixol in vivo: differential effect of the drug on brain oxidative status of scopolamine-treated rats

Several clinical studies implicated oxidative stress in the pathophysiology of both psychosis and dementia. As dementia is commonly associated with psychosis, antipsychotic medications are of importance in the pharmacotherapy of dementia particularly as a number of antipsychotics were reported to demonstrate neuronal pro-oxidant and/or antioxidant properties. Impairment of learning and memory, as the most characteristic manifestation of dementia, could be induced in experimental animals by acute administration of scopolamine (SCO) with a resultant elevation in brain oxidative status. This study investigated the potential pro-oxidant and/or antioxidant activity of the antipsychotic drug zuclopenthixol acetate, as its effect on brain oxidative status has yet to be evaluated. A 2 x 3 between-subjects factorial design was used to investigate the simultaneous and interactive effects of zuclopenthixol (0.7 and 1.4 mg/kg i.p.) and SCO on rat brain malondialdehyde, glutathione, glutathione peroxidase and superoxide dismutase levels/activities. Results revealed a significant pro-oxidant effect for both zuclopenthixol and SCO alone conditions. In addition, combined treatment of zuclopenthixol and SCO was found to be significantly different compared to either treatment conditions with regard to their effect on different brain oxidative stress indices. Such findings may have valuable implications in the pharmacotherapy of both psychosis and dementia.     

In vitro osteoclast resorption of bone substitute biomaterials used for implant site augmentation: a pilot study

PURPOSE: This observational study examined the resorptive behavior of normal neonatal rabbit osteoclasts grown on slices of bovine cortical bone as compared to samples of commercially available bone substitute biomaterials. It also examined the surface characteristics of these materials. MATERIALS AND METHODS: The 11 materials tested fell into 3 groups: (1) bone-derived, including freeze-dried human rib block, human demineralized freeze-dried bone, and deproteinated bovine bone; (2) synthetic hydroxyapatites (HA); and (3) synthetic non-HA, including coated methacrylates and coated silica glass. After 4 days in culture, 1 group of samples of each material underwent scanning electron microscopy (SEM) to evaluate resorptive pitting versus controls, while another group underwent tartrate-resistant acid phosphatase staining and light microscopy to examine osteoclast numbers and morphology. The 2 bovine-derived HA materials also underwent immunohistochemical staining and surface chemistry analysis. RESULTS: While most of these materials supported osteoclast attachment, some spreading, and survival in culture, only the bone-derived materials, with the exception of sintered deproteinated bovine bone, showed large scalloped-edged resorption pits with trails and exposed collagen when examined by SEM, although not to the same extent as unprocessed natural bone material. The HA materials and the sintered deproteinated bovine bone showed evidence of etching with smaller pits but no evidence of resorptive trail formation. The non-HA materials showed no evidence of pit formation or trails. Under immunohistochemical staining, Bio-Oss appeared to be positive for type I collagen after osteoclast activity on its surface, while Osteograf/N showed no positive staining. Surface chemistry analysis revealed nitrogen present in Bio-Oss specimens (0.17% to 0.47%), while there was no nitrogen detected in the Osteograf/N (0.00%); the percent nitrogen observed in normal bovine bone controls was 6.01% to 9.25%. DISCUSSION: The bone-derived materials supported osteoclast activity on the material surface in a way that facilitated formation of the more complex resorption pits in vitro. Assuming the rate of pit formation observed in vitro mimics that observed in vivo, the quantity and type of osteoclastic remodeling seen on non-bone-derived materials--and perhaps sintered bone-derived materials--would be extremely slow to negligible. Physiologic removal of non-bone-derived bone substitutes in vivo may occur by methods other than osteoclast resorption. CONCLUSIONS: Allogenous and xenogenous bone-derived materials that undergo delayed physiologic resorption may be more appropriately used with a staged surgical approach when used in sites intended to support osseointegrated dental implants. The combination of collagen staining and the presence of nitrogen suggest that there may be residual protein in Bio-Oss.     

Characteristics of long-term live-donor renal allograft survivors

BACKGROUND/AIMS: Despite the high rate of rejection, allograft failure and patient mortality in the early years of renal allotransplantation, some patients have done-remarkably well. We report here on 62 renal transplant recipients out of 144 patients (43%) who had functioning grafts for more than 15 years (range 15-24 years). MATERIALS: Demographic and follow-up data for patients fulfilling the criteria were reviewed. These patients include 43 males and 19 females, with a mean age at transplantation of 27.5 +/- 6.6 years (range 9-43 years), and mean donor age of 30 +/- 8.6 years. The donor source was 8 parents, 49 siblings and 5 unrelated. The main causes of end-stage renal disease were chronic pyelonephritis and chronic glomerulonephritis. Twenty-nine patients were treated with cyclosporine (CsA) while 33 patients were primarily immunosuppressed by steroids and azathioprine. RESULTS: Acute rejection episodes occurred in 40 patients (64.3%), out of them 19 patients experienced two or more acute rejection episodes. Univariate analysis showed that recipient and donor age, HLA-DR matching, pre- and post-transplant hypertension, ATN, delayed diuresis and chronic allograft nephropathy are significant risk factors; while recipient age, delayed diuresis and post-transplant hypertension were still significant by multivariate analysis. CONCLUSIONS: We concluded that renal transplantation, even in its earliest years and despite the numerous complications, has provided 15 or more years of near-normal life to patients with end-stage renal disease. Certain characteristics of long-term renal allograft survivors include young donor/recipient pairs, good DR matching with less pre- and post-transplantation prevalence of hypertension.