Effects of testosterone, estrogen and progesterone on TNF-α mediated cellular damage in rat arthritic synovial fibroblasts

Sexual dimorphism is a well-established phenomenon in rheumatoid arthritis, with women exhibiting higher disease severity. Understanding the role of sex hormones using in vivo animal models is limited due to the systemic effects as well as the difficulty in exploring different dose combinations of the hormones simultaneously. However, cell culture systems pose ideal systems for exploring different combinations and concentrations of the hormones simultaneously. In this study, the procedure for isolation of arthritic fibroblasts was standardized using a combination of collagenase and trypsin based on maximal yield and viability after employing different enzymatic disaggregation procedures. The cultured synovial fibroblasts from arthritic rats did not differ significantly from normal rat fibroblasts in terms of proliferation or secretion of inflammatory mediators. Stimulation of fibroblasts with TNF-α was standardized and TNF-α stimulated rat arthritic synovial fibroblasts exhibited an ideal in vitro system for screening antiinflammatory molecules. The effects of physiological and pharmacological concentrations of testosterone, estrogen and progesterone were studied on TNF-α induced cellular damage in rat arthritic synovial fibroblasts. The results showed that estrogen and testosterone exerted antiinflammatory effects on rat arthritic synovial fibroblasts at physiological and pharmacological concentrations. However, there was no significant difference in the effects between physiological and pharmacological concentrations. Progesterone independently did not show any protective effects. In combination with physiological concentrations of estrogen, progesterone abrogated estrogen's protective effect but it exhibited protection in combination with pharmacological concentrations of estrogen.     

A randomized prospective analysis of alteration of hemostatic function in patients receiving tranexamic acid and hydroxyethyl starch (130/0.4) undergoing off pump coronary artery bypass surgery

Postoperative hemorrhagic complications is still one of the major problems in cardiac surgeries. It may be caused by surgical issues, coagulopathy caused by the side effects of the intravenous fluids administered to produce plasma volume expansion such as hydroxyl ethyl starch (HES). In order to thwart this hemorrhagic issue, few agents are available. Fibrinolytic inhibitors like tranexamic acid (TA) may be effective modes to promote blood conservation; but the possible complications of thrombosis of coronary artery graft, precludes their generous use in coronary artery bypass graft surgery. The issue is a balance between agents that promote coagulation and those which oppose it. Therefore, in this study we have assessed the effects of concomitant use of HES and TA. Thromboelastogram (TEG) was used to assess the effect of the combination of HES and TA. With ethical committee approval and patient's consent, 100 consecutive patients were recruited for the study. Surgical and anesthetic techniques were standardized. Patients fulfilling our inclusion criteria were randomly allocated into 4 groups of 25 each. The patients in group A received 20 ml/kg of HES (130/0.4), 10 mg/kg of T.A over 30 minutes followed by infusion of 1 mg/kg/hr over the next 12 hrs. The patients in group B received Ringer's lactate + TA at same dose. The patients in the Group C received 20 ml/kg of HES. Group D patients received RL. Fluid therapy was goal directed. Total blood loss was assessed. Reaction time (r), α angle, maximum amplitude (MA) values of TEG were assessed at baseline, 12, 36 hrs. The possible perioperative myocardial infraction (MI) was assessed by electrocardiogram (ECG) and troponin T values at the baseline, postoperative day 1. Duration on ventilator, length of stay (LOS) in the intensive care unit (ICU) were also assessed. The demographical profile was similar among the groups. Use of HES increased blood loss significantly (P < 0.05). Concomitant use of TA reduced blood loss when used along with HES. r value was prolonged at 12 hours in all the groups and α angle was reduced at 12 hours in all the groups, where as MA value was reduced at 12 th hour in the HES group compared to the baseline and increased in TA + HES group. These findings were statistically significant. No significant change in Troponin T values/ECG, duration of ventilation and LOS ICU was observed. No adverse events was noticed in any of the four groups. HES (130/0.4) used at a dose of 20 ml/kg seems to produce coagulopathy causing increased blood loss perioperatively. Hemodilution produced by fluid therapy seems to produce Coagulopathy as observed by TEG parameters. Concomitant use of TA with HES appears to reverse these changes without causing any adverse effects in patients undergoing OPCAB surgery.     

In vitro evaluation of novel low-pressure spark plasma sintered HA–BG composite scaffolds for bone tissue engineering

The low-pressure spark plasma sintering (SPS) technique is adopted to fabricate hydroxyapatite–bioglass (HA–BG) scaffolds while maintaining the physical properties of both components, including their bulk and relative density and hardness. However, prior to their orthopaedic and dental applications, these scaffolds must be validated via pre-clinical assessments. In the present study, scaffolds with different ratios of HA : BG, namely, 100 : 0 (HB 0 S), 90 : 10 (HB 10 S), 80 : 20 (HB 20 S) and 70 : 30 (HB 30 S) were fabricated. These scaffolds were characterized by investigating their physicochemical properties (X-ray diffraction (XRD) and surface wettability), bioactivity in a simulated body fluid (SBF) (field emission scanning electron microscopy (FESEM), Fourier-transform infrared spectroscopy (FTIR) and calcium dissolution), antimicrobial properties, biocompatibility and osteoinduction of human bone marrow-derived mesenchymal stromal cells (hBMSCs) and human monocyte immune cell response. The XRD and surface wettability results confirmed no formation of undesirable phases and the enhanced surface hydrophilicity of the scaffolds, respectively. The bioactivity in SBF indicated the formation of bone-like apatite on the surface of the scaffolds, corresponding to an increase in BG%, which was confirmed through FTIR spectra and the increasing trend of calcium release in SBF. The scaffolds showed inhibition properties against Staphylococcus aureus and Staphylococcus epidermidis. The scanning electron microscopy (SEM) micrographs and Alamar Blue proliferation assay indicated the good attachment and significant proliferation, respectively, of hBMSCs on the scaffolds. Alizarin Red S staining confirmed that the scaffolds supported the mineralisation of hBMSCs. The osteogenic protein secretion (bone morphogenetic protein-2 (BMP2), type-I collagen (COL1) and osterix (OSX)) was significant on the HB 30 S-seeded hBMSCs when compared with that of HB 0 S. The monocyte migration was significantly halted in response to HA–BG-conditioned media when compared with the positive control (monocyte chemoattractant protein-1: MCP-1). In conclusion, the HB 30 S composite scaffold has a greater potential to substitute bone grafts in orthopaedic and dental applications.

HB 30 S composite scaffold inhibits Staphylococcus spp., supports the biocompatibility and osteogenic differentiation of hBMSCs and resists monocyte migration.     

Gamma Knife radiosurgery in the management of brainstem metastases

Background

Metastases to the brainstem portend a poor prognosis and present a challenge in clinical management. Surgical resection is rarely a viable option.

Methods

Post-treatment MRI scans of patients with brainstem metastases treated with radiosurgery were used to determine local control and disease progression. Median survival was calculated using Kaplan–Meier analysis. Univariate and multivariate analyses were performed using log-rank test and Cox proportional hazards model, respectively.

Results

Thirty-two consecutive patients with brainstem metastasis underwent Gamma Knife radiosurgery. Median age was 50 years. Median tumor volume was 0.71 cm³ and median tumor margin dose was 13 Gy. Seventeen of 32 patients received WBRT prior to stereotactic radiosurgery. Median survival was 5.2 months. There was a statistically significant difference in survival based on RTOG recursive partition analysis (RPA) class. Median survival of patients categorized as RPA class I was 19.2 months, RPA class II was 8.4 months, and RPA class III was 1.9 months. The overall local tumor control rate was 87.5%. There were no acute complications following stereotactic radiosurgery and no evidence of radiation necrosis noted on post-treatment MRI scans.

Conclusion

Stereotactic radiosurgery is an effective treatment for brainstem metastases and should be considered especially for patients with good performance status.     

Susceptible and protective associations of HLA DRB1*/DQB1* alleles and haplotypes with ischaemic stroke

Stroke has emerged as the second commonest cause of mortality worldwide and is a major public health problem. For the first time, we present here the association of human leucocyte antigen (HLA)‐DRB1*/DQB1* alleles and haplotypes with ischaemic stroke in South Indian patients. Ischaemic stroke (IS) cases and controls were genotyped for HLA‐DRB1*/DQB1* alleles by polymerase chain reaction sequence‐specific primers (PCR‐SSP) method. The frequencies of HLA class II alleles such as DRB1*04, DRB1*07, DRB1*11, DRB1*12, DRB1*13, DQB1*02 and DQB1*07 were high in IS patients than in the age‐ and gender‐matched controls, suggesting that the individuals with these alleles are susceptible to ischaemic stroke in South India. The frequencies of alleles such as DRB1*03, DRB1*10, DRB1*14, DQB1*04 and DQB1*05 were less in IS cases than in the controls, suggesting a protective association. Haplotypes DRB1*04‐DQB1*0301, DRB1*07‐DQB1*02, DRB1*07‐DQB1*0301, DRB1*11‐DQB1*0301 and DRB1*13‐DQB1*06 were found to be high in IS patients conferring susceptibility. The frequency of haplotype DRB1*10‐DQB1*05 was high in controls conferring protection. IS‐LVD and gender‐stratified analysis too confirmed these susceptible and protective associations. Thus, HLA‐DRB1*/DQB1* alleles and haplotypes strongly predispose South Indian population to ischaemic stroke. Further studies in different populations with large sample size or the meta‐analysis are needed to explain the exact mechanism of associations of HLA gene(s) with IS.     

Number needed to treat and costs per responder among biologic treatments for moderate-to-severe psoriasis: a network meta-analysis

Background: The clinical benefits of biologic therapies for moderate-to-severe psoriasis are well established, but wide variations exist in patient response.

Objectives: To determine the number needed to treat (NNT) to achieve a 75% and 90% reduction in the Psoriasis Area and Severity Index (PASI-75/90) with FDA-approved agents and evaluate the incremental cost per PASI-75 or PASI-90 responder.

Methods: The relative probabilities of achieving PASI-75 and PASI-90, as well as NNTs, were estimated using a network meta-analysis. Costs (2017 USD) included drug acquisition and administration. The incremental cost per PASI-75 or PASI-90 responder for each treatment was estimated for the clinical trial period, and annually.

Results: Compared with supportive care, the NNT to achieve PASI-75 was 1.18 for ixekizumab, 1.29 for secukinumab 300?mg, 1.37 for infliximab, 1.48 for adalimumab, 1.53 for secukinumab 150?mg, 1.58 for ustekinumab, 2.25 for etanercept, and 3.71 for apremilast. The one-year incremental cost per PASI-75 responder relative to supportive care was $59,830 for infliximab, $88,775 for secukinumab 300?mg, $91,837 for adalimumab, $95,898 for ixekizumab, $97,363 for ustekinumab, $105,131 for secukinumab 150?mg, $129,665 for apremilast, and $159,328 for etanercept. Results were similar for PASI-90.

Conclusion: The NNT and incremental cost per responder are meaningful ways to assess comparative effectiveness and cost effectiveness among psoriasis treatments.     

Quantitation and characterization of glutathionyl haemoglobin as an oxidative stress marker in chronic renal failure by mass spectrometry

OBJECTIVES: Glutathionyl haemoglobin (GS-Hb) belonging to the class of glutathionylated proteins has been investigated as a possible marker of oxidative stress in different chronic diseases. The purpose of this study was to examine whether glutathionyl haemoglobin can serve as an oxidative stress marker in non-diabetic chronic renal failure patients on different renal replacement therapies (RRT) through its quantitation, and characterization of the specific binding site of glutathione in haemoglobin molecule by mass spectrometric analysis. DESIGN AND METHODS: The study group consisted of non-diabetic chronic renal failure patients on renal replacement therapy (RRT): hemodialysis (HD), continuous ambulatory peritoneal dialysis (CAPD) and renal allograft transplant (Txp) patients. Haemoglobin samples of these subjects were analyzed by liquid chromatography electrospray ionization mass spectrometry for GS-Hb quantitation. Characterization of GS-Hb was done by tandem mass spectrometry. Levels of erythrocyte glutathione (GSH) and lipid peroxidation (as thiobarbituric acid reacting substances) were measured spectrophotometrically, while glycated haemoglobin (HbA1c) was measured by HPLC. RESULTS: GS-Hb levels were markedly elevated in the dialysis group and marginally in the transplant group as compared to the controls. GS-Hb levels correlated positively with lipid peroxidation and negatively with the erythrocyte glutathione levels in RRT groups indicating enhanced oxidative stress. De novo sequencing of the chymotryptic fragment of GS-Hb established that glutathione is attached to Cys-93 of the beta globin chain. Mass spectrometric quantitation of total glycated haemoglobin showed good agreement with HbA1c estimation by conventional HPLC method. CONCLUSIONS: Glutathionyl haemoglobin can serve as a clinical marker of oxidative stress in chronic debilitating therapies like RRT. Mass spectrometry provides a reliable analytical tool for quantitation and residue level characterization of different post-translational modifications of haemoglobin.