Osteoclastic inhibition: an action of nitric oxide not mediated by cyclic GMP.

The osteoclast is unique in its ability to resorb bone, and excessive osteoclastic activity has been implicated in osteoporosis, Paget disease of bone, rheumatoid arthritis, and the growth of metastases in bone. The activity of this cell is controlled by the main circulating inhibitor, calcitonin, in association with locally produced modulators. We show that nitric oxide (NO) may be an important member of the latter group. NO is produced by the vascular endothelium and nervous system and is involved in both neurotransmission and the regulation of blood pressure. However, our results show that the autocoid is also a potent inhibitor of osteoclast function. NO (30 microM) produced a decrease to approximately 50% of the original osteoclast spread area. Similar effects were also produced by 3-morpholinosydnonimine or sodium nitroprusside, reagents that spontaneously release NO. These shape changes were associated with a reduction of bone resorption after a 24-hr incubation of isolated osteoclasts on devitalized bone slices. NO is thought to act by stimulating guanylate cyclase, with a consequent increase in cyclic GMP, but a different mode of action is likely in the osteoclast since dibutyryl or 8-bromo cyclic GMP have no effect. It should be noted that calcitonin can produce similar changes in shape and activity but is associated with an increase in osteoclast intracellular calcium and cessation of membrane movement; neither of these is produced by NO, suggesting that its mode of action is different. The abundance of NO-producing endothelial cells in bone marrow and their proximity to osteoclasts suggests that marrow endothelial cells may play a physiological role in the regulation of osteoclastic activity.     

Microalbuminuria as a predictor of early glomerular injury in children with sickle cell disease

Objective : A cross sectional study was carried out to determine the prevalence of microalbuminuria in the pediatric patients with sickle cell disease.Methods : The study was carried out on 64 pediatric patients aged less than 14 years with documented HbSS, HbAS and HbS beta thalassemia, Microalbuminuria was estimated using single radial immuno diffusion technique. Majority of the study subjects were of HbSS type. 38.5% had symptoms for >2 years. 18.8% of the study population had significant microalbuminuria (19.2% of SS types and 18.8% of Hb AS types).Result : Microalbuminuria excretion was significantly more in patients >9 years of age as compared to young patients (p<0.05). Mean serum creatinine levels did not show any significant difference in the various study groups.Conclusion : Microalbuminuria estimation is a very important clinical marker of preclinical glomerular damage in patients with sickle cell disease. Its estimation would help in the early detection of such patients and prompt initiation of therapy.     

Evaluation of Preoperative Serum Levels of CA 125 and Expression of p53 in Ovarian Neoplasms: A Prospective Clinicopathological Study in a Tertiary Care Hospital

Objectives

To assess the preoperative serum levels of CA 125 with its diagnostic role and to evaluate the p53 expression in patients of primary ovarian neoplasms. We also wished to judge their relationship with other parameters like clinical staging and histopathologic tumor type.

Materials and Methods

The present study was conducted on 86 patients during the study period of 2.5 years. Preoperative CA 125 levels were evaluated by an automated immunoassay analyzer. p53 expression was judged immunohistochemically with pre-diluted monoclonal antibody. An objective scoring was done depending on distinct nuclear immunopositivity.

Results

Median value of preoperative CA 125 levels was 32 U/mL in benign surface epithelial-stromal tumors (BSEST), 53 U/mL in borderline surface epithelial-stromal tumors (BOT), 346 U/mL in malignant surface epithelial-stromal tumors (MSEST) and 560 U/mL in serous adenocarcinomas (SAC). Most of ovarian tumors were in the FIGO stage I (64 cases, 74.4%), but higher stages (II, III, IV) were observed mostly in MSESTs. SACs displayed the maximum p53 expression. Considering the cut-off value of more than 35 U/mL in CA 125 levels, the sensitivity to diagnose MSESTs was 94.7%. Preoperative CA 125 levels strongly and positively correlated with FIGO staging and p53 expression. Similarly p53 expression strongly and positively correlated with FIGO staging and histopathological categories.

Conclusion

Higher values of preoperative CA 125 levels and higher expression p53 are associated with MSESTs and BOTs especially of serous type. They strongly correlate with each other and with tumor stage. But there is no serum CA 125 concentration that can clearly differentiate benign and malignant ovarian masses.     

Effect of bone extracellular matrix synthesized in vitro on the osteoblastic differentiation of marrow stromal cells

Alternative materials for bone grafts are gaining greater importance in dentistry and orthopaedics, as the limitations of conventional methods become more apparent. We are investigating the generation of osteoinductive matrix in vitro by culturing cell/scaffold constructs for tissue engineering applications. The main strategy involves the use of a scaffold composed of titanium (Ti) fibers seeded with progenitor cells. In this study, we investigated the effect of extracellular matrix (ECM) laid down by osteoblastic cells on the differentiation of marrow stromal cells (MSCs) towards osteoblasts. Primary rat MSCs were harvested from bone marrow, cultured in dexamethasone containing medium and seeded directly onto the scaffolds. Constructs were grown in static culture for 12 days and then decellularized by rapid freeze-thaw cycling. Decellularized scaffolds were re-seeded with pre-cultured MSCs at a density of 2.5 x 10(5) cells/construct and osteogenicity was determined according to DNA, alkaline phosphatase, calcium and osteopontin analysis. DNA content was higher for cells grown on decellularized scaffolds with a maximum content of about 1.3 x 10(6) cells/construct. Calcium was deposited at a greater rate by cells grown on decellularized scaffolds than the constructs with only one seeding on day-16. The Ti/MSC constructs showed negligible calcium content by day-16, compared with 213.2 (+/- 13.6) microg/construct for the Ti/ECM/MSC constructs cultured without any osteogenic supplements after 16 days. These results indicate that bone-like ECM synthesized in vitro can enhance the osteoblastic differentiation of MSCs.     

A genome-wide search for non-UGT1A1 markers associated with unconjugated bilirubin level reveals significant association with a polymorphic marker near a gene of the nucleoporin family

Variants in the UGT1A1 gene and its promoter are known to determine levels of unconjugated bilirubin (UCB), but do not explain all cases of unconjugated hyperbilirubinemia. To discover associations with variants in genes other than UGT1A1, we undertook a genome-wide association study. We recruited 200 participants to cover the entire range of quantitative variation in UCB level. The data set -- after data curation, including analyses for population stratification and cryptic relatedness -- comprised genotypes at 512,349 SNP loci on 182 individuals. Quantitative trait locus (QTL) association analyses were performed, after adjusting the UCB level for effects of age, gender, and genotype at the dinucleotide (TA) insertion locus in UGT1A1 that is known to significantly modulate UCB level. A significant association of a polymorphic marker (rs2328136) near the NUP153 gene (which produces a 153 kDa nucleoporin) was obtained (p = 0.002, after multiple-testing correction). The frequency of the variant allele (A) at the rs2328136 locus in our study population is 40%, higher than most global populations. NUP153, whose product is a major regulatory factor in bidirectional transport of biomolecules across nucleus to cytosol, is associated with the transport of biliverdin reductase, which is important for bilirubin conjugation.     

Apigenin, a dietary flavonoid, sensitizes human T cells for activation-induced cell death by inhibiting PKB/Akt and NF-kappaB activation pathway

Resistance of T cells to activation-induced cell death (AICD) is associated with autoimmunity and lymphoproliferation. We found that apigenin (4',5,7-trihydroxyflavone), a non-mutagenic dietary flavonoid, augmented both extrinsic and intrinsic pathways of apoptosis in recurrently activated, but not in primarily stimulated, human blood CD4+ T cells. Apigenin potentiated AICD by inhibiting NF-kappaB activation and suppressing NF-kappaB-regulated anti-apoptotic molecules, cFLIP, Bcl-x(L), Mcl-1, XIAP and IAP, but not Bcl-2. Apigenin suppressed NF-kappaB translocation to nucleus and inhibited IkappaBalpha phosphorylation and degradation in response to TCR stimulation in reactivated peripheral blood CD4 T cells, as well as in leukemic Jurkat T cell lines. Among the pathways that lead to NF-kappaB activation upon TCR stimulation, apigenin selectively inhibited PI3K-PKB/Akt, but not PKC-theta activation in the human T cells, and synergized with a PI3K inhibitor to markedly augment AICD. Apigenin also suppressed expression of anti-apoptotic cyclooxygenase 2 (COX-2) protein in activated human T cells, but it did not affect activation of Erk MAPKinase. Thus, in chronically activated human T cells, relatively non-toxic apigenin can suppress anti-apoptotic pathways involving NF-kappaB activation, and especially cFLIP and COX-2 expression that are important for functioning and maintenance of immune cells in inflammation, autoimmunity and lymphoproliferation.     

The long-term effectiveness of generic adult fixed-dose combination antiretroviral therapy for HIV-infected Ugandan children

Background

Access to pediatric antiretroviral formulations is increasing in resource-limited countries, however adult FDCs are still commonly used by antiretroviral therapy (ART) programs.

Objective

To describe long-term effectiveness of using adult FDC of d4T+3TC+NVP (Triomune) in children for HIV treatment.

Methods

Clinical, immunologic, and virologic outcomes of HIV-infected ART-naïve children aged six months to 12 years, were evaluated up to 96 weeks post-ART initiation.

Results

From March 2004 to June 2006, 104 children were followed with a median age of 5.4 years, median CD4 cell percent and HIV-1 RNA were 11.0% (IQR 6.7–13.9) and 348,846copies/mL (IQR 160,941–681,313) respectively at baseline. Using Kaplan-Meir estimates, 75% of children had undetectable viral loads (<400copies/mL) at 96weeks of ART. Children with a baseline CD4 cell percent >15% were 3 times more likely to achieve viral load <400copies/mL than those with baseline CD4 cell percent <5% after adjusting for baseline age {aHR = 3.03 (1.10–8.32), p=0.03}; no difference was found among those with CD4 cell percent >5–14.9% and <5%.

Conclusion

Treatment with generic adult FDC for HIV-infected Ugandan children led to sustained clinical, immunologic and virologic response during 96 weeks of ART. Early initiation of ART is key to achieving virological success.     

Fluoroscopic Caudal Epidural Injections in Managing Post Lumbar Surgery Syndrome: Two-Year Results of a Randomized,Double-Blind,Active-Control Trial

Study Design: A randomized, active control, double-blind trial. Objective: To evaluate the effectiveness of fluoroscopically directed caudal epidural injections with or without steroids in managing chronic low back and lower extremity pain secondary to post lumbar surgery syndrome. Summary of Background Data: There is a paucity of evidence concerning caudal epidural injections for managing chronic persistent low back pain with or without lower extremity pain caused by post lumbar surgery syndrome.Methods: This active control randomized study included 140 patients with 70 patients in each group. Group I received 0.5% lidocaine, 10 mL; Group II received 9 mL of 0.5% lidocaine mixed with 1 mL of 6 mg of nonparticulate betamethasone. The multiple outcome measures included the numeric rating scale, the Oswestry Disability Index 2.0, employment status, and opioid intake with assessments at 3, 6, 12, 18, and 24 months posttreatment. Primary outcome was defined as at least 50% improvement in pain and Oswestry Disability Index scores. Patients with a positive response to the first 2 procedures with at least 3 weeks of relief were considered to be successful. All others were considered as failures.Results: Overall in Group I, 53% and 47% of the patients and in Group II, 59% and 58% of the patients, showed significant improvement with reduction in pain scores and disability index at 12 months and 24 months. In contrast, in the successful groups, significant pain relief and improvement in function were observed in 70% and 62% of Group I at one and 2 years; in 75% and 69% of Group II at one and 2 years. The results in the successful group showed that at the end of the first year patients experienced approximately 38 weeks of relief and at the end of 2 years Group I had 62 weeks and Group II had 68 weeks of relief. Overall total relief for 2 years was 48 weeks in Group I and 54 weeks in Group II. The average procedures in the successful groups were at 4 in one year and 6 at the end of 2 years.Conclusion: Caudal epidural injections of local anesthetic with or without steroid might be effective in patients with chronic persistent low back and/or lower extremity pain in patients with post lumbar surgery syndrome.     

Mycobacterium bovis BCG Causing Vertebral Osteomyelitis (Pott’s Disease) Following Intravesical BCG Therapy

We report a case of Mycobacterium bovis BCG vertebral osteomyelitis in a 79-year-old man 2.5 years after intravesical BCG therapy for bladder cancer. The recovered isolate resembled M. tuberculosis biochemically, but resistance to pyrazinamide (PZA) rendered that diagnosis suspect. High-pressure liquid chromatographic studies confirmed the diagnosis of M. bovis BCG infection. The patient was originally started on a four-drug antituberculous regimen of isoniazid, rifampin, ethambutol, and PZA. When susceptibility studies were reported, the regimen was changed to isoniazid and rifampin for 12 months. Subsequently, the patient was transferred to a skilled nursing facility for 3 months, where he underwent intensive physical therapy. Although extravesical adverse reactions are rare, clinicians and clinical microbiologists need to be aware of the possibility of disseminated infection by M. bovis BCG in the appropriate setting of clinical history, physical examination, and laboratory investigation.     

Variations in the functional domain of basal core promoter of hepatitis B virus among Eastern Indian patients with prevalence of genotypes A, C, and D among the same ethnic population

Mutations in the basal core promoter (BCP) and precore (PC) regions are associated with persistent and intermittently high hepatitis B virus (HBV) replication in several patients. The variability in the functional domains of BCP and PC region of HBV and their association with disease progression and clinical outcome were assessed in Eastern India, an unique region where three HBV genotypes, A, D, and C are prevalent among the same ethnic group. PCR amplification and direct sequencing of BCP and PC region was done on sera obtained from 130 HBsAg positive subjects with different clinical presentations. Associations of the apparent risk factors with clinical advancement were evaluated by statistical methods including multiple logistic regression analyses (MLR). HBV genotype A was present in 33.08%, C in 25.38%, and D in 41.54% cases. Genotypes A and C were associated with higher rate of T1762/A1764 mutations than the most predominant genotype D. HBeAg negative state was associated with considerably higher rate of C1753 mutation. T1762/A1764 along with C1753 was common among cirrhosis and T1762/A1764 without C1753 was frequent among chronic liver disease cases. No significant association was found between A1896 point mutation and clinical status. Multivariate analysis revealed that T1762/A1764 double mutation, HBV/A, age ≥25 years, C1753 and A1899 were critical factors for clinical advancement while age ≥25 years and C1753 as significant predictor for cirrhosis in comparison with chronic liver disease. In conclusion, the analysis of the BCP variability may help in monitoring the progression towards advanced liver disease in Eastern Indian patients.