Does hand stiffness reflect internal organ fibrosis in diabetes mellitus?

Fibrosis leads to irreversible stiffening of tissue and loss of function, and is a common pathway leading to morbidity and mortality in chronic disease. Diabetes mellitus (both type 1 and type 2 diabetes) are associated with significant fibrosis in internal organs, chiefly the kidney and heart, but also lung, liver and adipose tissue. Diabetes is also associated with the diabetic cheirarthropathies, a collection of clinical manifestations affecting the hand that include limited joint mobility (LJM), flexor tenosynovitis, Duypuytren disease and carpal tunnel syndrome. Histo-morphologically these are profibrotic conditions affecting various soft tissue components in the hand. We hypothesize that these hand manifestations reflect a systemic profibrotic state, and are potential clinical biomarkers of current or future internal organ fibrosis. Epidemiologically, there is evidence that fibrosis in one organ associates with fibrosis with another; the putative exposures that lead to fibrosis in diabetes (advanced glycation end product deposition, microvascular disease and hypoxia, persistent innate inflammation) are 'systemic'; a common genetic susceptibility to fibrosis has also been hinted at. These data suggest that a subset of the diabetic population is susceptible to multi-organ fibrosis. The hand is an attractive biomarker to clinically detect this susceptibility, owing to its accessibility to physical examination and exposure to repeated mechanical stresses. Testing the hypothesis has a few pre-requisites: being able to measure hand fibrosis in the hand, using clinical scores or imaging based scores, which will facilitate looking for associations with internal organ fibrosis using validated methodologies for each. Longitudinal studies would be essential in delineating fibrosis trajectories in those with hand manifestations. Since therapies reversing fibrosis are few, the onus lies on identification of a susceptible subset for preventative measures. If systematically validated, clinical hand examination could provide a low-cost, universally accessible and easily reproducible screening step in selecting patients for clinical trials for fibrosis in diabetes.

IgA Determines Bacterial Composition in the Gut.

Background: Classically, IgA in the gut prevents the invasion of microorganisms to systemic organs through the process of neutralization and immune exclusion. Interestingly, recent reports suggest that IgA might help in biofilm formation and promote bacterial growth inside the intestine. Methods: In this study, we used flow cytometry, ELISA, and chemical models of colitis to test whether the quality and quantity of IgA can select for bacterial persistence in the gut. Results: We found that members of Proteobacteria, such as γ-Proteobacteria and SFB, are preferentially coated by IgA in WT mice. In the partial absence of either T-dependent or -independent IgA responses, there are no significant differences in the frequency of bacteria coated with IgA in mice. However, Rag-/- mice that lack all antibodies had a severe reduction in Proteobacteria and were resistant to DSS-induced colitis, suggesting that secretory IgA might be essential for differential retention of these taxa in the mouse gut. Rag-/- littermates in the F2 generation generated from (B6 × Rag-/-) F1 mice acquired the underrepresented bacteria taxa such as γ-Proteobacteria through vertical transmission of flora. They died soon after weaning, possibly due to the acquired flora. Additionally, continued exposure of Rag-/- mice to B6 flora by cohousing mice led to the acquisition of γ-Proteobacteria and mortality. Conclusions: Together, our results indicate that host survival in the complete absence of an IgA response necessitates the exclusion of specific bacterial taxa from the gut microbiome.

Differential Regulation of Two Arms of mTORC1 Pathway Fine-Tunes Global Protein Synthesis in Resting B Lymphocytes.

Protein synthesis is tightly regulated by both gene-specific and global mechanisms to match the metabolic and proliferative demands of the cell. While the regulation of global protein synthesis in response to mitogen or stress signals is relatively well understood in multiple experimental systems, how different cell types fine-tune their basal protein synthesis rate is not known. In a previous study, we showed that resting B and T lymphocytes exhibit dramatic differences in their metabolic profile, with implications for their post-activation function. Here, we show that resting B cells, despite being quiescent, exhibit increased protein synthesis in vivo as well as ex vivo. The increased protein synthesis in B cells is driven by mTORC1, which exhibits an intermediate level of activation in these cells when compared with resting T cells and activated B cells. A comparative analysis of the transcriptome and translatome of these cells indicates that the genes encoding the MHC Class II molecules and their chaperone CD74 are highly translated in B cells. These data suggest that the translatome of B cells shows enrichment for genes associated with antigen processing and presentation. Even though the B cells exhibit higher mTORC1 levels, they prevent the translational activation of TOP mRNAs, which are mostly constituted by ribosomal proteins and other translation factors, by upregulating 4EBP1 levels. This mechanism may keep the protein synthesis machinery under check while enabling higher levels of translation in B cells.

Transient systemic inflammation in adult male mice results in underweight progeny.

PROBLEM: While the testes represent an immune-privileged organ, there is evidence that systemic inflammation is accompanied by local inflammatory responses. We therefore examined whether transient systemic inflammation caused any inflammatory and functional consequences in murine testes. METHOD OF STUDY: Using a single systemic administration of Toll-like receptor (TLR) agonists [lipopolysaccharide (LPS) or peptidoglycan (PG) or polyinosinic-polycytidylic acid (polyIC)] in young adult male mice, we assessed testicular immune-inflammatory landscape and reproductive functionality. RESULTS: Our findings demonstrated a significant induction of testicular TNF-α, IL-1ß and IL-6 transcripts within 24 h of TLR agonist injection. By day 6, these cytokine levels returned to baseline. While there was no change in caudal sperm counts at early time points, eight weeks later, twofold decrease in sperm count and reduced testicular testosterone levels were evident. When these mice were subjected to mating studies, no differences in mating efficiencies or litter sizes were observed compared with controls. Nonetheless, the neonatal weights of progeny from LPS/PG/polyIC-treated sires were significantly lower than controls. Postnatal weight gain up to three weeks was also slower in the progeny of LPS/polyIC-treated sires. Placental weights at 17.5 days post-coitum were significantly lower in females mated to LPS- and polyIC-treated males. Given this likelihood of an epigenetic effect, we found lower testicular levels of histone methyltransferase enzyme, mixed-lineage leukaemia-1, in mice given LPS/PG/polyIC 8 weeks earlier. CONCLUSION: Exposure to transient systemic inflammation leads to transient local inflammation in the testes, with persistent sperm-mediated consequences for foetal development.

DBCOVP: A database of coronavirus virulent glycoproteins.

Since the emergence of SARS-CoV-1 (2002), novel coronaviruses have emerged periodically like the MERS- CoV (2012) and now, the SARS-CoV-2 outbreak which has posed a global threat to public health. Although, this is the third zoonotic coronavirus breakout within the last two decades, there are only a few platforms that provide information about coronavirus genomes. None of them is specific for the virulence glycoproteins and complete sequence-structural features of these virulence factors across the betacoronavirus family including SARS-CoV-2 strains are lacking. Against this backdrop, we present DBCOVP (http://covp.immt.res.in/), the first manually-curated, web-based resource to provide extensive information on the complete repertoire of structural virulent glycoproteins from coronavirus genomes belonging to betacoronavirus genera. The database provides various sequence-structural properties in which users can browse and analyze information in different ways. Furthermore, many conserved T-cell and B-cell epitopes predicted for each protein are present that may perform a significant role in eliciting the humoral and cellular immune response. The tertiary structure of the epitopes together with the docked epitope-HLA binding-complex is made available to facilitate further analysis. DBCOVP presents an easy-to-use interface with in-built tools for similarity search, cross-genome comparison, phylogenetic, and multiple sequence alignment. DBCOVP will certainly be an important resource for experimental biologists engaged in coronavirus research studies and will aid in vaccine development.

Febrile temperature change modulates CD4 T cell differentiation via a TRPV channel-regulated Notch-dependent pathway.

Fever is a conserved and prominent response to infection. Yet, the issue of how CD4 T cell responses are modulated if they occur at fever temperatures remains poorly addressed. We have examined the priming of naive CD4 T cells in vitro at fever temperatures, and we report notable fever-mediated modulation of their cytokine commitment. When naive CD4 T cells were primed by plate-bound anti-CD3 and anti-CD28 monoclonal antibodies at moderate fever temperature (39 °C), they enhanced commitment to IL4/5/13 (Th2) and away from IFNg (Th1). This was accompanied by up-regulation of the Th2-relevant transcription factor GATA3 and reduction in the Th1-relevant transcription factor Tbet. Fever sensing by CD4 T cells involved transient receptor potential vanilloid cation channels (TRPVs) since TRPV1/TRPV4 antagonism blocked the febrile Th2 switch, while TRPV1 agonists mediated a Th2 switch at 37 °C. The febrile Th2 switch was IL4 independent, but a γ-secretase inhibitor abrogated it, and it was not found in Notch1-null CD4 T cells, identifying the Notch pathway as a major mediator. However, when naive CD4 T cells were primed via antigen and dendritic cells (DCs) at fever temperatures, the Th2 switch was abrogated via increased production of IL12 from DCs at fever temperatures. Thus, immune cells directly sense fever temperatures with likely complex physiological consequences.

A role for cell-autocrine interleukin-2 in regulatory T-cell homeostasis.

Activated T-cells make both interleukin-2 (IL2) and its high-affinity receptor component CD25. Regulatory CD4 T-cells (Treg cells) do not make IL2, and the IL2-CD25 circuit is considered a paracrine circuit crucial in their generation and maintenance. Yet, all T-cells are capable of making IL2 at some stage during differentiation, making a cell-intrinsic autocrine circuit additionally possible. When we re-visited experiments with mixed bone marrow chimeras using a wide range of ratios of wild-type (WT) and IL2-/- genotype progenitors, we found that, as expected, thymic Treg cells were almost equivalent between WT and IL2-/- genotypes at ratios with WT prominence. However, at WT-limiting ratios, the IL2-/- genotype showed lower thymic Treg frequencies, indicating a role for cell-intrinsic autocrine IL2 in thymic Treg generation under IL2-limiting conditions. Further, peripheral IL2-/- naive CD4 T-cells showed poor conversion to inducible Tregs (pTregs) both in vivo and in vitro, again indicating a significant role for cell-intrinsic autocrine IL2 in their generation. Peripherally, the IL2-/- genotype was less prominent at all WT:IL2-/- ratios among both thymic Tregs (tTregs) and pTregs, adoptively transferred IL2-/- Tregs showed poorer survival than WT Tregs did, and RNA-seq analysis of WT and IL2-/- Tregs showed interesting differences in the T-cell receptor and transforming growth factor-beta-bone morphogenetic protein-JNK pathways between them, suggesting a non-titrating role for cell-intrinsic autocrine IL2 in Treg programming. These data indicate that cell-intrinsic autocrine IL2 plays significant roles in Treg generation and maintenance.

dEMBF v2.0: An Updated Database of Enzymes for Microalgal Biofuel Feedstock.

In light of increasing algal genomics data and knowledge of biosynthetic pathways responsible for biofuel production, an integrated resource for easy access to all information is essential to improve our understanding of algal lipid metabolism. Against this backdrop, dEMBF v2.0, a significantly updated and improved version of our database of microalgae lipid biosynthetic enzymes for biofuel production, has been developed. dEMBF v2.0 now contains a comprehensive annotation of 2018 sequences encoding 35 enzymes, an increase of over 7-fold as compared with the first version. Other improved features include an increase in species coverage to 32 algal genomes, analysis of additional metabolic pathways, expanded annotation thoroughly detailing sequence and structural features, including enzyme-ligand interactions, and integration of supporting experimental evidence to demonstrate the role of enzymes in increasing lipid content. Along with a complete redesign of the interface, the updated database provides several inbuilt tools and user-friendly functionalities for more interactive and dynamic visualization of data.

Characterization of biological variation of peripheral blood immune cytome in an Indian cohort.

Immune parameters show characteristic normal baseline levels and variance in the population. We characterised the degree of inter-individual and within-individual variation over one-year time period in 33 immune cell subsets by flow cytometry in peripheral blood mononuclear cells from 43 healthy young adult volunteers. Our analysis revealed that immune subsets that showed low variability between individuals also showed low short-term fluctuations within-individuals, as well as concordance in siblings. However, when baseline levels and degree of fluctuation were considered together, individuals failed to cluster into discreet groups. Together, the data reveal complex inter-relationships between immune subsets in individuals, and provide insights into the observed heterogeneity between individuals and between multiple immune subsets.

Role of NF-kappaB2-p100 in regulatory T cell homeostasis and activation.

The immunological roles of the nuclear factor-kappaB (NF-κB) pathway are mediated via the canonical components in immune responses and via non-canonical components in immune organogenesis and homeostasis, although the two components are capable of crosstalk. Regulatory CD4 T cells (Tregs) are homeostatically functional and represent an interesting potential meeting point of these two NF-κB components. We show that mice deficient in the non-canonical NF-κB component gene Nfkb2 (p100) had normal thymic development and suppressive function of Tregs. However, they had enhanced frequencies of peripheral 'effector-phenotype' Tregs (eTregs). In bi-parental chimeras of wild-type (WT) and Nfkb2-/- mice, the Nfkb2-/- genotype was over-represented in Tregs, with a further increase in the relative prominence of eTregs. Consistent with distinct properties of eTregs, the Nfkb2-/- genotype was more prominent in Tregs in extra-lymphoid tissues such as liver in the bi-parental chimeras. The Nfkb2-/- Tregs also displayed greater survival, activation and proliferation in vivo. These Nfkb2-/- Tregs showed higher nuclear NF-κB activity mainly comprising of RelB-containing dimers, in contrast to the prominence of cRel- and RelA-containing dimers in WT Tregs. Since p100 is an inhibitor of RelB activation as well as a participant as cleaved p52 in RelB nuclear activity, we tested bi-parental chimeras of WT and Relb-/- mice, and found normal frequencies of Relb-/- Tregs and eTregs in these chimeric mice. Our findings confirm and extend recent data, and indicate that p100 normally restrains RelB-mediated Treg activation, and in the absence of p100, p50-RelB dimers can contribute to Treg activation.

Functionally significant metabolic differences between B and T lymphocyte lineages.

Activation of B and T lymphocytes leads to major remodelling of the metabolic landscape of the cells enabling their post-activation functions. However, naive B and T lymphocytes also show metabolic differences, and the genesis, nature and functional significance of these differences are not yet well understood. Here we show that resting B-cells appeared to have lower energy demands than resting T-cells as they consumed lower levels of glucose and fatty acids and produced less ATP. Resting B-cells are more dependent on OXPHOS, while T-cells show more dependence on aerobic glycolysis. However, despite an apparently higher energy demand, T lineage cells showed lower rates of protein synthesis than equivalent B lineage stages. These metabolic differences between the two lineages were established early during lineage differentiation, and were functionally significant. Higher levels of protein synthesis in B-cells were associated with increased synthesis of MHC class II molecules and other proteins associated with antigen internalization, transport and presentation. The combination of higher energy demand and lower protein synthesis in T-cells was consistent with their higher ATP-dependent motility. Our data provide an integrated perspective of the metabolic differences and their functional implications between the B and T lymphocyte lineages.

Clinical and Immunological Profile of Anti-factor H Antibody Associated Atypical Hemolytic Uremic Syndrome: A Nationwide Database.

Background: Atypical hemolytic uremic syndrome (aHUS), an important cause of acute kidney injury (AKI), is characterized by dysregulation of the alternative complement pathway. Autoantibodies to factor H (FH), a chief regulator of this pathway, account for a distinct subgroup. While high anti-FH titers predict relapse, they do not correlate well with disease activity and their functional characterization is required. Methods: Of 781 patients <18-year-old of aHUS in the nationwide database from 2007 to 2018, 436 (55.8%) had anti-FH antibodies. Clinical features and outcome of patients managed in the last 6-year (n = 317) were compared to before (n = 119). In plasma samples of 44 patients, levels of serial circulating FH immune complexes (CIC), free FH, soluble terminal complement complex (sC5b-9), sheep red blood cell (SRBC) lysis and epitope specificity (n = 8) were examined. Functional renal reserve, ambulatory hypertension, left ventricular hypertrophy (LVH), and proteinuria were evaluated in a subset. Results: Patients presented with markedly elevated anti-FH titers (10,633.2 ± 998.5 AU/ml). Management varied by center, comprising plasma exchange (PEX; 77.5%) and immunosuppression (73.9%). Patients managed in the last 6-year showed better renal survival at mean 28.5 ± 27.3 months (log rank P = 0.022). Mean anti-FH titers stayed 700-1,164 AU/ml during prolonged follow-up, correlating with CIC. Patients with relapse had lower free-FH during remission [Generalized estimating equations (GEE), P = 0.001]; anti-FH levels ≥1,330 AU/ml and free FH ≤440 mg/l predicted relapse (hazards ratio, HR 6.3; P = 0.018). Epitope specificity was similar during onset, remission and relapse. Antibody titer ≥8,000 AU/ml (HR 2.23; P = 0.024), time to PEX ≥14 days (HR 2.09; P = 0.071) and PEX for <14 days (HR 2.60; P = 0.017) predicted adverse renal outcomes. Combined PEX and immunosuppression improved long-term outcomes (HR 0.37; P = 0.026); maintenance therapy reduced risk of relapses (HR 0.11; P < 0.001). At 4.4±2.5 year, median renal reserve was 15.9%; severe ambulatory, masked and pre-hypertension were found in 38, 30, and 18%, respectively. Proteinuria and LVH occurred in 58 and 28% patients, respectively. Conclusion: Prompt recognition and therapy with PEX and immunosuppression, is associated with satisfactory outcomes. Free-FH predicts early relapses in patients with high anti-FH titers. A significant proportion of impaired functional reserve, ambulatory hypertension, proteinuria and LVH highlight the need for vigilant long-term follow-up.

Gut IgA abundance in adult life is a major determinant of resistance to dextran sodium sulfate-colitis and can compensate for the effects of inadequate maternal IgA received by neonates.

Studies with gene-deficient and gnotobiotic mice have identified many host and microbial factors that contribute to induced colitis, but information on whether specific factors determine susceptibility under more physiological conditions is lacking. Using wild-type strains that differ in their IgA response but harbor a diverse gut microbiome, we found that the IgA-high strain CBA/CaJ (CBA) is resistant to acute colitis induced with dextran sodium sulfate (DSS), unlike the IgA-low strain C57BL/6 (B6). Resistance was associated with extensive IgA-coating of fecal bacteria, lower fecal bacterial loads and greater abundance of barrier-protective transcripts in colonic tissues under homeostatic conditions. Fecal microbial transplant (FT) experiments revealed that disease induction in B6 mice was associated with a cohort of bacteria that are not targeted by IgA. However, CBA mice continued to be resistant to colitis induction following FTs from B6 mice, indicating that they are able to contain such colitogenic members. In support of a role for bacterial exclusion in resistance, oral administration of immunoglobulins decreased DSS-induced disease in B6 mice. In F1 mice derived separately with CBA and B6 dams and in F1 mice backcrossed to the two parental strains, resistance segregated with the IgA response of the pups and not with barrier-associated transcripts or bacterial loads. Interestingly, B6 pups foster-nursed on CBA dams continued to be susceptible in later life, whereas CBA pups foster-nursed on B6 dams continued to be resistant. Together, the data indicate that a high-IgA response in adult life can protect against colitis and compensate for IgA deficiency in early life.

Altered Peripheral Blood Leucocyte Phenotype and Responses in Healthy Individuals with Homozygous Deletion of FHR1 and FHR3 Genes.

A homozygous 83-kb deletion encompassing the genes for complement factor-H-related proteins 1 and 3 (FHR 1, FHR3) is known as a risk factor for some immune inflammatory disorders. However, the functional relevance of this FHR1/3 deletion is relatively unexplored. Globally, healthy populations of all ethnic groups tested show an 8-10% prevalence of homozygosity for this deletion polymorphism. We have begun to compare the peripheral leucocyte phenotype and functionality between FHR1/3-/- and FHR1/3+/+ healthy adult individuals. We report that the two groups show significant differences in their peripheral blood innate leucocyte subset composition, although the adaptive immune subsets are similar between them. Specifically, FHR1/3-/- individuals show higher frequencies of patrolling monocytes and lower frequencies of classical monocytes than FHR1/3+/+ individuals. Similarly, FHR1/3-/- individuals show higher frequencies of plasmacytoid dendritic cells (pDCs) and lower frequencies of myeloid DCs (mDCs) than FHR1/3+/+ individuals. Notably, classical monocytes specifically showed cell-surface-associated factor H (FH), and cells from the FHR1/3-/- group had somewhat higher surface-associated FH levels than those from FHR1/3+/+ individuals. FHR1/3-/- monocytes also showed elevated secretion of TNF-α, IL-1ß, and IL-10 in response to TLR7/8 or TLR4 ligands. Similarly, FHR1/3-/- mDCs and pDCs showed modest but evident hyper-responsiveness to TLR ligands. Our findings, that the FHR1/3-/- genotype is associated with significant alterations of both the relative prominence and the functioning of monocyte and DC subsets, may be relevant in understanding the mechanism underlying the association of the genotype with immune inflammatory disorders.

Correlation of cell-surface CD8 levels with function, phenotype and transcriptome of naive CD8 T cells.

We have previously demonstrated co-receptor level-associated functional heterogeneity in apparently homogeneous naive peripheral CD4 T cells, dependent on MHC-mediated tonic signals. Maturation pathways can differ between naive CD4 and naive CD8 cells, so we tested whether the latter showed similar co-receptor level-associated functional heterogeneity. We report that, when either polyclonal and T-cell receptor (TCR)-transgenic monoclonal peripheral naive CD8 T cells from young mice were separated into CD8hi and CD8lo subsets, CD8lo cells responded poorly, but CD8hi and CD8lo subsets of CD8 single-positive (SP) thymocytes responded similarly. CD8lo naive CD8 T cells were smaller and showed lower levels of some cell-surface molecules, but higher levels of the negative regulator CD5. In addition to the expected peripheral decline in CD8 levels on transferred naive CD8 T cells in wild-type (WT) but not in MHC class I-deficient recipient mice, short-duration naive T-cell-dendritic cell (DC) co-cultures in vitro also caused co-receptor down-modulation in CD8 T cells but not in CD4 T cells. Constitutive pZAP70/pSyk and pERK levels ex vivo were lower in CD8lo naive CD8 T cells and dual-specific phosphatase inhibition partially rescued their hypo-responsiveness. Bulk mRNA sequencing showed major differences in the transcriptional landscapes of CD8hi and CD8lo naive CD8 T cells. CD8hi naive CD8 T cells showed enrichment of genes involved in positive regulation of cell cycle and survival. Our data show that naive CD8 T cells show major differences in their signaling, transcriptional and functional landscapes associated with subtly altered CD8 levels, consistent with the possibility of peripheral cellular aging.

Cell-intrinsic regulation of peripheral memory-phenotype T cell frequencies.

Memory T and B lymphocyte numbers are thought to be regulated by recent and cumulative microbial exposures. We report here that memory-phenotype lymphocyte frequencies in B, CD4 and CD8 T-cells in 3-monthly serial bleeds from healthy young adult humans were relatively stable over a 1-year period, while Plasmablast frequencies were not, suggesting that recent environmental exposures affected steady state levels of recently activated but not of memory lymphocyte subsets. Frequencies of memory B and CD4 T cells were not correlated, suggesting that variation in them was unlikely to be determined by cumulative antigenic exposures. Immunophenotyping of adult siblings showed high concordance in memory, but not of recently activated lymphocyte subsets. To explore the possibility of cell-intrinsic regulation of T cell memory, we screened effector memory-phenotype T cell (TEM) frequencies in common independent inbred mice strains. Using two pairs from these strains that differed predominantly in either CD4 TEM and/or CD8 TEM frequencies, we constructed bi-parental bone marrow chimeras in F1 recipient mice, and found that memory T cell frequencies in recipient mice were determined by donor genotypes. Together, these data suggest cell-autonomous determination of memory T niche size, and suggest mechanisms maintaining immune variability.

Differences in multiple immune parameters between Indian and U.S. infants.

To compare immune phenotypes across two geographic and ethnic communities, we examined umbilical cord blood by flow cytometry and Luminex in parallel cohorts of 53 newborns from New Delhi, India, and 46 newborns from Stanford, California. We found that frequencies of a B cell subset suggested to be B-1-like, and serum IgM concentration were both significantly higher in the Stanford cohort, independent of differences in maternal age. While serum IgA levels were also significantly higher in the Stanford cohort, IgG1, IgG2, and IgG4 were significantly higher in the New Delhi samples. We found that neutrophils, plasmacytoid dendritic cells, CD8+ T cells, and total T cells were higher in the U.S. cohort, while dendritic cells, patrolling monocytes (CD14dimCD16+), natural killer cells, CD4+ T cells, and naïve B cells were higher in the India cohort. Within the India cohort, we also identified cell types whose frequency was positively or negatively predictive of occurrence of infection(s) in the first six months of life. Monocytes, total T cells, and memory CD4+ T cells were most prominent in having an inverse relationship with infection. We suggest that these data provide impetus for follow-up studies linking phenotypic differences to environmental versus genetic factors, and to infection outcomes.

Opioid-free anesthesia for breast cancer surgery: An observational study.

BACKGROUND AND AIMS: Opioids are associated with postoperative nausea, vomiting, drowsiness, and increased analgesic requirement. A nonopioid anesthesia technique may reduce morbidity, enable day care surgery, and possibly decrease tumor recurrence. We compared opioid-free, nerve block-based anesthesia with opioid-based general anesthesia for breast cancer surgery in a prospective cohort study. MATERIAL AND METHODS: Twenty four adult American Society of Anesthesiologists grade I-III patients posted for modified radical mastectomy (MRM) with axillary dissection were induced with propofol and maintained on isoflurane (0.8-1.0 minimum alveolar concentration) through i-gel on spontaneous ventilation and administered ultrasound-guided PECS 1 and 2 blocks (0.1% lignocaine + 0.25% bupivacaine + 1 mcg/kg dexmedetomidine, 30 ml). Postoperative nausea, pain scores, nonopioid analgesic requirement over 24 h, stay in the recovery room, and satisfaction of surgeon and patient were studied. Twenty-four patients who underwent MRM and axillary dissection without a nerve block under routine opioid anesthesia with controlled ventilation were the controls. RESULTS: MRM and axillary dissection under the nonopioid technique was adequate in all patients. Time in the recovery room, postoperative nausea, analgesic requirement, and visual analog scale scores were all significantly less in the nonopioid group. Surgeon and patient were satisfied with good patient quality of life on day 7. CONCLUSION: Nonopioid nerve block technique is adequate and safe for MRM with axillary clearance. Compared to conventional technique, it offers lesser morbidity and may allow for earlier discharge. Larger studies are needed to assess the long-term impact on chronic pain and tumor recurrence by nonopioid techniques.

Characterization of genetic predisposition and autoantibody profile in atypical haemolytic-uraemic syndrome.

We previously reported that Indian paediatric patients with atypical haemolytic-uraemic syndrome (aHUS) showed high frequencies of anti-complement factor H (FH) autoantibodies that are correlated with homozygous deletion of the genes for FH-related proteins 1 and 3 (FHR1 and FHR3) (FHR1/3-/- ). We now report that Indian paediatric aHUS patients without anti-FH autoantibodies also showed modestly higher frequencies of the FHR1/3-/- genotype. Further, when we characterized epitope specificities and binding avidities of anti-FH autoantibodies in aHUS patients, most anti-FH autoantibodies were directed towards the FH cell-surface anchoring polyanionic binding site-containing C-terminal short conservative regions (SCRs) 17-20 with higher binding avidities than for native FH. FH SCR17-20-binding anti-FH autoantibodies also bound the other cell-surface anchoring polyanionic binding site-containing region FH SCR5-8, at lower binding avidities. Anti-FH autoantibody avidities correlated with antibody titres. These anti-FH autoantibody characteristics did not differ between aHUS patients with or without the FHR1/3-/- genotype. Our data suggest a complex matrix of interactions between FHR1-FHR3 deletion, immunomodulation and anti-FH autoantibodies in the aetiopathogenesis of aHUS.

Interaction of CD80 with Neph1: a potential mechanism of podocyte injury.

BACKGROUND: The induction of CD80 on podocytes has been shown in animal models of podocyte injury and in certain cases of nephrotic syndrome. In a lipopolysaccharide (LPS)-induced mouse model of albuminuria, we have recently shown a signalling axis of LPS-myeloid cell activation-TNFα production-podocyte CD80 induction-albuminuria. Therefore, in this report, we investigated the cellular and molecular consequences of TNFα addition and CD80 expression on cultured podocytes. METHODS: A murine podocyte cell line was used for TNFα treatment and for over-expressing CD80. Expression and localization of various podocyte proteins was analysed by reverse transcriptase-polymerase chain reaction, western blotting and immunofluorescence. HEK293 cells were used to biochemically characterize interactions. RESULTS: Podocytes treated with LPS in vitro did not cause CD80 upregulation but TNFα treatment was associated with an increase in CD80 levels, actin derangement and poor wound healing. Podocytes stably expressing CD80 showed actin derangement and co-localization with Neph1. CD80 and Neph1 interaction was confirmed by pull down assays of CD80 and Neph1 transfected in HEK293 cells. CONCLUSION: Addition of TNFα to podocytes causes CD80 upregulation, actin reorganization and podocyte injury. Overexpressed CD80 and Neph1 interact via their extracellular domain. This interaction implies a mechanism of slit diaphragm disruption and possible use of small molecules that disrupt CD80-Neph1 interaction as a potential for treatment of nephrotic syndrome associated with CD80 upregulation.