Spectrum of renal vascular lesions among patients with lupus nephritis: An experience from a tertiary care center.

Background: Lupus nephritis (LN) is the assemblage of glomerular, tubulointerstitial and vascular changes. Despite the fact that glomerular changes are overemphasized in pathological classification and scoring system, but the existence of vascular damage negatively impact the clinical course. Aims and Objective: This study was conducted to determine the clinicopathological spectrum of renal vascular lesions in lupus nephritis. Materials and Methods: Renal microvascular lesions in biopsy proven lupus nephritis were classified into 5 major categories-thrombotic microangiopathy, true vasculitis; lupus vasculopathy, uncomplicated vascular immune deposits, and arterial. Clinical details, laboratory parameters and histopathological variables were compared among all groups. Summary of chronic changes was also assessed. Results: Biopsies from 56 patients revealed thrombotic microangiopathy (2), lupus vasculopathy (3), uncomplicated vascular immune deposit (6), PAN type vasculitis (1) and arterial sclerosis (13). No renal vascular lesions were found in 35.18% of patients. At the time of biopsy, arterial sclerosis or lupus vasculopathy patients were older Nephritis subtype. Activity indices were higher in lupus vasculopathy group whereas patients with arteriosclerosis showed highest chronicity index. Conclusions: Renal vascular lesions are common in systemic lupus erythematosus patients with nephritis and may be associated with aggressive clinical course.

mMCP7, a Mouse Ortholog of δ Tryptase, Mediates Pelvic Tactile Allodynia in a Model of Chronic Pelvic Pain.

Chronic prostatitis/Chronic pelvic pain syndrome (CP/CPPS) is a condition that affects a large number of men and has unknown etiology. We have previously demonstrated the presence of elevated levels of mast cell tryptase in expressed prostatic secretions (EPS) of CP/CPPS patients. In a murine model of CP/CPPS, we showed tryptase and its cognate receptor PAR2 as critical to the development of pelvic pain and lower urinary tract symptoms. Here, we extend these observations to demonstrate that an isoform of tryptase called delta (δ)-tryptase, is elevated in the EPS of patients with CP/CPPS and is correlated with pelvic pain symptoms. Using an Escherichia coli (CP1) -induced murine model of CP/CPPS, we demonstrated a differential response in C57BL/6J and NOD/ShiLtJ mice, with C57BL6/J mice being resistant to an increase in pelvic tactile allodynia, despite having equivalent levels of activated mast cells in the prostate. Activated tryptase+ve mast cells were observed to be in closer apposition to PGP9.5+ve nerve fibers in the prostate stroma of NOD/ShiLtJ in comparison to C57BL/6J mice. The mouse ortholog of δ-tryptase, mouse mast cell protease 7 (mMCP7) has been reported to be unexpressed in C57BL/6J mice. We confirmed the absence of mMCP7 in the prostates of C57BL/6J and its presence in NOD/ShiLtJ mice. To evaluate a role for mMCP7 in the differential allodynia responses, we performed direct intra-urethral instillations of mMCP7 and the beta (ß)-tryptase isoform ortholog, mMCP6 in the CP1-infection model. mMCP7, but not mMCP6 was able to induce an acute pelvic allodynia response in C57BL/6J mice. In-vitro studies with mMCP7 on cultured mast cells as well as dissociated primary neurons demonstrated the ability to induce differential activation of pain and inflammation associated molecules compared to mMCP6. We conclude that mMCP7, and possibility its human ortholog δ-tryptase, may play an important role in mediating the development of pelvic tactile allodynia in the mouse model of pelvic pain and in patients with CP/CPPS.

Post encroachment time threshold identification for right-turn related crashes at unsignalized intersections on intercity highways under mixed traffic.

Unsignalized intersections in developing countries experience many serious conflicts between cross-traffic due to indiscipline traffic manoeuvrability. Historically, Post Encroachment Time (PET) has gained attention as a proximal indicator to analyze crossing conflicts. However, identifying an appropriate PET threshold to classify critical conflicts for highly heterogeneous traffic scenario is still an unexplored area. Consequently, this study proposes a novel approach of PET threshold identification with proof of application by collecting data from ten intersections located on four-lane intercity highways in the National Capital Region (NCR), India. Both crossing conflicts and right-turn related crash data (for the left-hand drive) are collected. Their correlations are thoroughly studied for each PET threshold using a quantitative technique considering all and individual vehicle categories. Finally, a qualitative analysis is done by ranking the sites based on cumulative PET and related crashes to verify the proposed quantitative technique. A PET threshold of 1 s is obtained from both the techniques which can be used to identify critical conflicts for unsignalized intersections located on four-lane intercity highways. The proposed methodology will serve as an alternative, faster and effective tool to evaluate the hazardousness of unsignalized intersections located on intercity highways under highly heterogeneous traffic condition.

Protease-activated receptor 2 activates CRAC-mediated Ca2+ influx to cause prostate smooth muscle contraction.

Protease activated receptor 2 (PAR2) is a G-protein coupled receptor that contributes to prostate fibrosis and lower urinary tract symptoms (LUTS). In addition to fibrosis, aberrant smooth muscle tone in the prostate has been hypothesized to play a role. We therefore examined PAR2 expression in primary human prostate smooth muscle cells (PSMC) and studied the downstream signaling effects of PAR2 activation. Signaling pathways involved in the process were assessed using the PAR2 activating peptide SLIGKV-NH2. We show that PAR2 is expressed in PSMC and that PAR2 activation mediates a biphasic elevation in intracellular Ca2+ and phosphorylation of myosin light chain 20 (MLC20), causing cellular contraction as assessed in a gel contraction assay. Intracellular Ca2+ flux was inhibited by a phosphoinositide hydrolysis inhibitor, U73122, showing a requirement for phospholipase C ß (PLCß) activation. PSMC expressed mRNA for L-type voltage dependent Ca2+ channels (VDCC) as well as Ca2+ release activated channels (CRAC), a hitherto unreported finding. Secondary intracellular Ca2+ oscillations were abrogated only by BTP2, the CRAC channel inhibitor, but not by nifedipine, an inhibitor of VDCC. These data suggest that, PAR2 activation and subsequent Ca2+ entry through CRAC channels are important mechanisms in prostate smooth muscle contraction.

Synemin: Molecular Features and the Use of Proximity Ligation Assay to Study Its Interactions.

Synemin has three splice variants (α, ß, and L) with identical head and rod domains but with tail domains of varying size. α- and ß-Synemin are larger than most intermediate filament proteins (1565 and 1253 amino acids, respectively) but L-synemin is shorter (339 amino acids). Synemin isoforms do not self-assemble into filaments but can copolymerize with vimentin and desmin. Synemin is present in all muscle cell types, in a few neural cell types, and in various other nonepithelial cell types. Synemin expression is regulated, sometimes in an isoform-specific manner, during development of the nervous system, in brain and breast cancer cells and during injuries to the brain and liver. Mice-lacking synemin develop a myopathic phenotype, possibly due to synemin role in linking desmin filaments to costameres and sarcomeres. Synemin may play this role through its demonstrated binding to costameric and sarcolemmal proteins, such as α-actinin, vinculin, and members of the dystroglycan complex. In astrocytoma cells, synemin regulates proliferation by interacting with PP2A to modulate Akt phosphorylation status. Methods to identify synemin binding partners are central to understand the roles of this protein in diverse cell types. Here, we describe how to use proximal ligation assays (PLA) for this purpose. PLA complement biochemical methods such as immunoprecipitation by relying on the use of antibodies conjugated to oligonucleotide probes to visualize by fluorescence microscopy protein-protein interactions in cells and tissues.

Roles and potential clinical applications of intermediate filament proteins in brain tumors.

Intermediate filament (IF) proteins are cytoplasmic and nuclear cytoskeletal proteins. Of the ~70 IF proteins, nearly 12 are found in the nervous system, where their expression is largely cell-type specific. Astrocytes express glial fibrillary acidic protein (GFAP), whereas different neuron types contain neurofilament proteins, α-internexin, or peripherin. These proteins are often downregulated in brain cancer. In addition, brain cancer cells may also contain vimentin, nestin, and synemin, which are the IF proteins found in neural progenitor cells. In different brain tumor types, the expression of nestin, vimentin, and α-internexin appears to correlate with the clinical outcome. Experimental investigations have also demonstrated that IF proteins have distinct roles in specific brain tumor cell behaviors: nestin, for instance, is important for the proliferation of glioma cells, whereas synemin also affect their mobility. The mechanisms responsible for these effects involve the interaction of IF proteins with specific signaling pathways. Synemin, for instance, positively regulates glioma cell proliferation by antagonizing protein phosphatase 2A. Further evidence for the potential of IF proteins as therapeutic targets derives from animal models showing the influence of IF proteins on tumor growth. Nestin downregulation, for instance, dramatically reduced intracerebral glioma growth. Selective targeted therapies of IFs to date primarily include gene therapy approaches using nestin or GFAP gene promoters to drive transgene expression into glioma cells. Although attempts to identify small molecules specifically antagonizing IF proteins have been unsuccessful to date, it is anticipated that the identification of such compounds will be instrumental in expanding therapeutic approaches for brain tumors.

Synemin promotes AKT-dependent glioblastoma cell proliferation by antagonizing PP2A.

The intermediate filament protein synemin is present in astrocyte progenitors and glioblastoma cells but not in mature astrocytes. Here we demonstrate a role for synemin in enhancing glioblastoma cell proliferation and clonogenic survival, as synemin RNA interference decreased both behaviors by inducing G1 arrest along with Rb hypophosphorylation and increased protein levels of the G1/S inhibitors p21(Cip1) and p27(Kip1). Akt involvement was demonstrated by decreased phosphorylation of its substrate, p21(Cip1), and reduced Akt catalytic activity and phosphorylation at essential activation sites. Synemin silencing, however, did not affect the activities of PDPK1 and mTOR complex 2, which directly phosphorylate Akt activation sites, but instead enhanced the activity of the major regulator of Akt dephosphorylation, protein phosphatase type 2A (PP2A). This was accompanied by changes in PP2A subcellular distribution resulting in increased physical interactions between PP2A and Akt, as shown by proximity ligation assays (PLAs). PLAs and immunoprecipitation experiments further revealed that synemin and PP2A form a protein complex. In addition, treatment of synemin-silenced cells with the PP2A inhibitor cantharidic acid resulted in proliferation and pAkt and pRb levels similar to those of controls. Collectively these results indicate that synemin positively regulates glioblastoma cell proliferation by helping sequester PP2A away from Akt, thereby favoring Akt activation.

Impact assessment and recommendation of alternative conjunctive water use strategies for salt affected agricultural lands through a field scale decision support system - a case study.

Conjunctive use of saline/non-saline irrigation waters is generally aimed at minimizing yield losses and enhancing flexibility of cropping, without much alteration in farming operations. Recommendation of location-specific suitable conjunctive water use plans requires assessment of their long-term impacts on soil salinization/sodification and crop yield reductions. This is conventionally achieved through long-term field experiments. However such impact evaluations are site specific, expensive and time consuming. Appropriate decision support systems (DSS) can be time-efficient and cost-effective means for such long-term impact evaluations. This study demonstrates the application of one such (indigenously developed) DSS for recommending best conjunctive water use plans for a, rice-wheat growing, salt affected farmer's field in Gurgaon district of Haryana (India). Before application, the DSS was extensively validated on several farmers and controlled experimental fields in Gurgaon and Karnal districts of Haryana (India). Validation of DSS showed its potential to give realistic estimates of root zone soil salinity (with R = 0.76-0.94; AMRE = 0.03-0.06; RMSPD = 0.51-0.90); sodicity (with R = 0.99; AMRE = 0.02; RMSPD = 0.84) and relative crop yield reductions (AMRE = 0.24), under existing (local) resource management practices. Long term (10 years) root zone salt build ups and associated rice/wheat crop yield reductions, in a salt affected farmer's field, under varied conjunctive water use scenarios were evaluated with the validated DSS. It was observed that long-term applications of canal (CW) and tube well (TW) waters in a cycle and in 1:1 mixed mode, during Kharif season, predicted higher average root zone salt reductions (2-9%) and lower rice crop yield reductions (4-5%) than the existing practice of 3-CW, 3-TW, 3-CW. Besides this, long-term application of 75% CW mixed with 25% TW, during Rabi season, predicted about 17% lower average root-zone salt reductions than the cyclic applications of (1-CW, 1-TW, 2-CW) and (2-CW, 1-TW, 1-CW, i.e., existing irrigation strategy). However, average wheat crop yield reductions (16-17%) simulated under all these strategies were almost at par. In general, cyclic-conjunctive water use strategies emerged as better options than the blending modes. These results were in complete confirmation with actual long-term conjunctive water use experiments on similar soils. It was thus observed that such pre-validated tools could be efficient means for designing, local resource and target crop yield-specific, appropriate conjunctive water use plans for irrigated agricultural lands.