Outcomes of Management of Severe Clubfoot among Children by Ilizarov External Fixator.

Introduction: Clubfoot remains the most common birth defect involving the musculoskeletal system. There are various surgical and non-surgical treatment options available for the management of clubfoot. Using the minimally invasive Ilizarov external fixator method has been reported to have good success rates and fewer complications. Materials and methods: This study aimed at analysing the morphological and functional outcomes of treating severe clubfoot by Ilizarov external fixator among children from July 2017 to March 2020. Thirty-two children who had either failed Ponseti / surgery or neglected with 44 clubfeet of Dieglio type III and type IV were included in the study. A short-leg walking cast was applied for an additional six weeks after removing of Ilizarov frame and additionally followed by an orthosis for another six weeks. Outcomes were measured by the functional rating system by Laaveg and Ponseti and interpretation done at 1 month and 12 months after the ankle-foot arthrosis. Results: About 86.4% of the patients had good or excellent outcome scores. Pre and post-Demeglio scores and functional rating scores were statistically significant (p<0.001) by using Paired t-test. Complications included superficial pin site infections in 13 feet (29.54%), 5 feet (11.36%) had claw toes, 3 feet (6.81%) had linear skin necrosis and 2 feet (4.54%) had calcaneal fractures which were manageable with minor interventions. Conclusion: The study findings highlighted that the Ilizarov external fixator method can correct complex foot deformities of severe clubfoot with minimum morbidity. Further larger and long-term studies are needed to investigate the effects of the stiff hindfoot and possible degenerative changes on the function and symptoms of these patients as adults.

Interferon-ε is a tumour suppressor and restricts ovarian cancer.

High-grade serous ovarian cancers have low survival rates because of their late presentation with extensive peritoneal metastases and frequent chemoresistance1, and require new treatments guided by novel insights into pathogenesis. Here we describe the intrinsic tumour-suppressive activities of interferon-ε (IFNε). IFNε is constitutively expressed in epithelial cells of the fallopian tube, the cell of origin of high-grade serous ovarian cancers, and is then lost during development of these tumours. We characterize its anti-tumour activity in several preclinical models: ovarian cancer patient-derived xenografts, orthotopic and disseminated syngeneic models, and tumour cell lines with or without mutations in Trp53 and Brca genes. We use manipulation of the IFNε receptor IFNAR1 in different cell compartments, differential exposure status to IFNε and global measures of IFN signalling to show that the mechanism of the anti-tumour activity of IFNε involves direct action on tumour cells and, crucially, activation of anti-tumour immunity. IFNε activated anti-tumour T and natural killer cells and prevented the accumulation and activation of myeloid-derived suppressor cells and regulatory T cells. Thus, we demonstrate that IFNε is an intrinsic tumour suppressor in the female reproductive tract whose activities in models of established and advanced ovarian cancer, distinct from other type I IFNs, are compelling indications of potential new therapeutic approaches for ovarian cancer.

Characterization of Monoclonal Antibodies to Measure Cell Surface Protein Levels of Human Interferon-Lambda Receptor 1.

Type III interferons (IFN-lambdas, IFN-λs) are important antiviral cytokines that can also modulate immune responses by acting through a heterodimeric receptor composed of the specific and limited expressed IFN-λR1 chain and the ubiquitous IL-10R2 chain, which is shared with IL-10 family cytokines. Conflicting data have been reported regarding which cells express the IFN-λR1 subunit and directly respond to IFN-λs. This is, in part, owing to transcript levels of the IFN-λR1 gene, IFNLR1, not always correlating with cell surface protein levels. In this study, we tested a panel of novel monoclonal antibodies (mAbs) that specifically recognize human IFN-λR1. Initially, antigen specificity was confirmed by enzyme-linked immunosorbent assay (ELISA), from which a subset of antibodies was selected for additional flow cytometry and neutralization assays. We further characterized two antibodies based on their strong ELISA binding activity (HLR1 and HLR14) and found only HLR14 could reliably detect cell surface IFN-λR1 protein on a variety of cell lines by flow cytometry. HLR14 could also detect IFN-λR1 protein on certain primary human blood cells, including plasmacytoid dendritic cells and B cells from peripheral blood. Availability of the HLR14 mAb will enable the quantification of IFN-λR1 protein levels on cells and better characterization of the cell specificity of the IFN-λ response.

Constitutive expression and distinct properties of IFN-epsilon protect the female reproductive tract from Zika virus infection.

The immunological surveillance factors controlling vulnerability of the female reproductive tract (FRT) to sexually transmitted viral infections are not well understood. Interferon-epsilon (IFNɛ) is a distinct, immunoregulatory type-I IFN that is constitutively expressed by FRT epithelium and is not induced by pathogens like other antiviral IFNs α, ß and λ. We show the necessity of IFNɛ for Zika Virus (ZIKV) protection by: increased susceptibility of IFNɛ-/- mice; their "rescue" by intravaginal recombinant IFNɛ treatment and blockade of protective endogenous IFNɛ by neutralising antibody. Complementary studies in human FRT cell lines showed IFNɛ had potent anti-ZIKV activity, associated with transcriptome responses similar to IFNλ but lacking the proinflammatory gene signature of IFNα. IFNɛ activated STAT1/2 pathways similar to IFNα and λ that were inhibited by ZIKV-encoded non-structural (NS) proteins, but not if IFNε exposure preceded infection. This scenario is provided by the constitutive expression of endogenous IFNε. However, the IFNɛ expression was not inhibited by ZIKV NS proteins despite their ability to antagonise the expression of IFNß or λ. Thus, the constitutive expression of IFNɛ provides cellular resistance to viral strategies of antagonism and maximises the antiviral activity of the FRT. These results show that the unique spatiotemporal properties of IFNε provides an innate immune surveillance network in the FRT that is a significant barrier to viral infection with important implications for prevention and therapy.

Outcomes of Management of Severe Clubfoot among Children by Ilizarov External Fixator

@#Introduction: Clubfoot remains the most common birth defect involving the musculoskeletal system. There are various surgical and non-surgical treatment options available for the management of clubfoot. Using the minimally invasive Ilizarov external fixator method has been reported to have good success rates and fewer complications. Materials and methods: This study aimed at analysing the morphological and functional outcomes of treating severe clubfoot by Ilizarov external fixator among children from July 2017 to March 2020. Thirty-two children who had either failed Ponseti / surgery or neglected with 44 clubfeet of Dieglio type III and type IV were included in the study. A short-leg walking cast was applied for an additional six weeks after removing of Ilizarov frame and additionally followed by an orthosis for another six weeks. Outcomes were measured by the functional rating system by Laaveg and Ponseti and interpretation done at 1 month and 12 months after the ankle-foot arthrosis. Results: About 86.4% of the patients had good or excellent outcome scores. Pre and post-Demeglio scores and functional rating scores were statistically significant (p<0.001) by using Paired t-test. Complications included superficial pin site infections in 13 feet (29.54%), 5 feet (11.36%) had claw toes, 3 feet (6.81%) had linear skin necrosis and 2 feet (4.54%) had calcaneal fractures which were manageable with minor interventions. Conclusion: The study findings highlighted that the Ilizarov external fixator method can correct complex foot deformities of severe clubfoot with minimum morbidity. Further larger and long-term studies are needed to investigate the effects of the stiff hindfoot and possible degenerative changes on the function and symptoms of these patients as adults.

Spatiotemporal regulation of human IFN-ε and innate immunity in the female reproductive tract.

Although published studies have demonstrated that IFN-ε has a crucial role in regulating protective immunity in the mouse female reproductive tract, expression and regulation of IFN-ε in the human female reproductive tract (hFRT) have not been characterized to our knowledge. We obtained hFRT samples from a well-characterized cohort of women to enable us to comprehensively assess ex vivo IFN-ε expression in the hFRT at various stages of the menstrual cycle. We found that among the various types of IFNs, IFN-ε was uniquely, selectively, and constitutively expressed in the hFRT epithelium. It had distinct expression patterns in the surface and glandular epithelia of the upper hFRT compared with basal layers of the stratified squamous epithelia of the lower hFRT. There was cyclical variation of IFN-ε expression in the endometrial epithelium of the upper hFRT and not in the distal FRT, consistent with selective endometrial expression of the progesterone receptor and regulation of the IFNE promoter by progesterone. Because we showed IFN-ε stimulated important protective IFN-regulated genes in FRT epithelium, this characterization is a key element in understanding the mechanisms of hormonal control of mucosal immunity.

Proteomic analysis shows that the main constituent of subepidermal localised cutaneous amyloidosis is not galectin-7.

Lichen or macular localised cutaneous amyloidoses have long been described as keratinic amyloidoses and believed to be due to the deposition of cytokeratin peptides originating from epidermis in the dermal papillae. However, recently it was suggested that galectin-7 is the causative protein for this type of amyloidosis. This was based on the detection of galectin-7 in a biopsy from a patient diagnosed with Bowen's disease and localised cutaneous amyloidosis. In this study we report mass spectrometry-based proteomic analysis of the protein composition of localised cutaneous amyloid deposits from seven patients using laser microdissection and show that basal keratins are the main constituents of the amyloid deposits. Galectin-7 was not present in the dermal amyloid deposits and was only present in the overlying Congo red negative epidermis.

Mass Spectrometry-Based Method Targeting Ig Variable Regions for Assessment of Minimal Residual Disease in Multiple Myeloma.

Multiple myeloma is a systemic malignancy of monoclonal plasma cells that accounts for 10% of hematologic cancers. With development of highly effective therapies for multiple myeloma, minimal residual disease (MRD) assessment has emerged as an important end point for management decisions. Currently, serologic assays lack the sensitivity for MRD assessment, and invasive bone marrow sampling with flow cytometry or molecular methods has emerged as the gold standard. We report a sensitive and robust targeted mass spectrometry proteomics method to detect MRD in serum, without the need of invasive, sequential bone marrow aspirates. The method detects Ig-derived clonotypic tryptic peptides predicted by sequencing the clonal plasma cell Ig genes. A heavy isotope-labeled Ig internal standard is added to patient serum at a known concentration, the Ig is enriched in a light chain type specific manner, and proteins are digested and analyzed by targeted mass spectrometry. Peptides from the constant regions of the λ or κ light chains, Ig heavy chains, and clonotypic peptides unique to the patient monoclonal Igs are targeted. This technique is highly sensitive and specific for the patient-specific monoclonal Igs, even in samples negative by multiparametric flow cytometry. Our method can accurately and precisely detect monoclonal protein in serum of patients treated for myeloma and has broad implications for management of hematologic patients.

Structural integrity with functional plasticity: what type I IFN receptor polymorphisms reveal.

The type I IFNs activate an array of signaling pathways, which are initiated after IFNs bind their cognate receptors, IFNα/ß receptor (IFNAR)1 and IFNAR2. These signals contribute to many aspects of human health including defense against pathogens, cancer immunosurveillance, and regulation of inflammation. How these cytokines interact with their receptors influences the quality of these signals. As such, the integrity of receptor structure is pivotal to maintaining human health and the response to immune stimuli. This review brings together genome wide association studies and clinical reports describing the association of nonsynonymous IFNAR1 and IFNAR2 polymorphisms with clinical disease, including altered susceptibility to viral and bacterial pathogens, autoimmune diseases, cancer, and adverse reactions to live-attenuated vaccines. We describe the amino acid substitutions or truncations induced by these polymorphisms and, using the knowledge of IFNAR conformational changes, IFNAR-IFN interfaces and overall structure-function relationship of the signaling complexes, we hypothesize the effect of these polymorphisms on receptor structure. That these predicted changes to IFNAR structure are associated with clinical manifestations of human disease, highlights the importance of IFNAR structural integrity to maintaining functional quality of these receptor-mediated responses. Type I IFNs are pivotal to innate immune responses and ultimately, to human health. Understanding the consequences of altered structure on the actions of these clinically significant cell receptors provides important information on the roles of IFNARs in health and disease.

PROPERTIES AND FUNCTIONS OF THE NOVEL TYPE I INTERFERON EPSILON.

Interferon epsilon (IFNε) is a type I IFN with unusual patterns of expression and therefore, function. It is constitutively expressed by reproductive tract epithelium and regulated by hormones during estrus cycle, reproduction, and menopause and by exogenous hormones. The IFNe protein is encoded by a gene in the type I IFN locus, binds to IFNAR1 and 2 which are required for signaling via the JAK STAT pathway. Its affinity for binding receptors and transducing signals is less potent than IFNα or ß subtypes in vitro. Nevertheless, in vivo experiments indicate its efficacy in regulating mucosal immune responses and protecting from bacterial and viral infections. These studies demonstrate a different mechanism of action to type I IFNs. In this organ system with dynamic fluxes in cellularity, requirement to tolerate an implanted fetus, and be protected from disease, there is co-option of a special IFN from a family of effective immunoregulators, with unique controls and modified potency to make it a safe and effective constitutive reproductive tract cytokine.

Using Cellulolytic Nitrogen Fixing Bacterium, Azomonas agilis for Effective Degradation of Agricultural Residues.

INTRODUCTION: Azomonas agilis, a nitrogen-fixing bacterium, was isolated from rhizospheric soil in central Myanmar. METHODS & MATERIALS: The nitrogen-fixing activity of this bacterium was detected by plate screening method using glucose nitrogen free mineral medium and ammonium test-kit Cellulolytic activity was screened by plat assay and detected by Dinitrosalicyclic acid method (DNS). RESULTS & DISCUSSION: The isolated A. agilis grew in media containing 3-12% of NaCl, although the growth became poor when NaCl concentrations increased. Among various carbon sources, sucrose was the best source for ammonium accumulation of this bacterium, whereas arabinose was not the suitable carbon source. Although the nitrogen-fixing activity of A. agilis was highest after one week incubation, cellulase enzyme production was highest after 2-3 days of incubation. It was observed that cellulase enzyme activity of A. agilis for cellulose and sodium carboxymethyl cellulose (CMC) was almost the same. Three agricultural wastes were used to detect the cellulase enzyme activity of A. agilis, cellulase activity was better on filter paper as a substrate when compared to rice-straw and sawdust. CONCLUSION: So, the isolated A. agilis has high potential as an effective bacterial strain to use in sustainable agriculture and degradation of some agricultural residues.

A hot spot on interferon α/ß receptor subunit 1 (IFNAR1) underpins its interaction with interferon-ß and dictates signaling.

The interaction of IFN-ß with its receptor IFNAR1 (interferon α/ß receptor subunit 1) is vital for host-protective anti-viral and anti-proliferative responses, but signaling via this interaction can be detrimental if dysregulated. Whereas it is established that IFNAR1 is an essential component of the IFNAR signaling complex, the key residues underpinning the IFN-ß-IFNAR1 interaction are unknown. Guided by the crystal structure of the IFN-ß-IFNAR1 complex, we used truncation variants and site-directed mutagenesis to investigate domains and residues enabling complexation of IFN-ß to IFNAR1. We have identified an interface on IFNAR1-subdomain-3 that is differentially utilized by IFN-ß and IFN-α for signal transduction. We used surface plasmon resonance and cell-based assays to investigate this important IFN-ß binding interface that is centered on IFNAR1 residues Tyr240 and Tyr274 binding the C and N termini of the B and C helices of IFN-ß, respectively. Using IFNAR1 and IFN-ß variants, we show that this interface contributes significantly to the affinity of IFN-ß for IFNAR1, its ability to activate STAT1, the expression of interferon stimulated genes, and ultimately to the anti-viral and anti-proliferative properties of IFN-ß. These results identify a key interface created by IFNAR1 residues Tyr240 and Tyr274 interacting with IFN-ß residues Phe63, Leu64, Glu77, Thr78, Val81, and Arg82 that underlie IFN-ß-IFNAR1-mediated signaling and biological processes.

Radiotherapy, chemotherapy and atherosclerosis.

PURPOSE OF REVIEW: This article reviews the effects of radiotherapy and chemotherapy in promoting the progression of atherosclerosis in patients with cancer. RECENT FINDINGS: Radiotherapy is associated with an increase in the incidence of atherosclerosis with the effects being related to the site of irradiation and dose of radiotherapy. Cranial irradiation is associated with dyslipidaemia and the metabolic syndrome secondary to effect on growth hormone secretion. Chemotherapeutic oncological therapies are associated with numerous cardiac diseases including valve disease, pericarditis and cardiomyopathy but can also promote atherosclerosis. Therapies directed against vascular endothelial growth factor including tyrosine kinase inhibitors have a direct effect in raising blood pressure and increase rates of cardiovascular disease (CVD) events. Antimetabolites such as 5-fluorouraciland capecitabine cause chest pain and increase CVD events. Anthracyclines cause heart failure and may increase CVD risk. SUMMARY: There is increasing evidence that radiotherapy and some chemotherapeutic agents are associated with increased rates of CVD. Patients who have received treatment for cancer should be considered at higher risk of developing atherosclerosis and require increased monitoring, further investigation and earlier treatment than would be suggested by classical risk factor management strategies.

Interferon epsilon promotes HIV restriction at multiple steps of viral replication.

Interferon epsilon (IFNɛ) is a type I IFN that is expressed constitutively in the female reproductive tract (FRT), and contributes to protection in models of sexually transmitted infections. Using multiple cell systems, including reporter cell lines and activated peripheral blood lymphocytes (PBLs), we show that recombinant IFNɛ impairs HIV infection at stage(s) post HIV entry and up to the translation of viral proteins. Consistent with this, IFNɛ upregulated a number of host cell restriction factors that block HIV at these stages of the replication cycle. The potency of IFNɛ induction of these HIV restriction factors was comparable to conventional type I IFNs, namely IFNα and IFNß. IFNɛ also significantly reduced the infectivity of progeny virion particles likely by inducing expression of HIV restriction factors, such as IFITM3, which act at that stage of infection. Thus, our data demonstrate that human IFNɛ suppresses HIV replication at multiple stages of infection.

Influenza A Virus in Backyard Pigs and Poultry in Rural Cambodia.

Surveillance of influenza virus in humans and livestock is critical, given the worldwide public health threats and livestock production losses. Livestock farming involving close proximity between humans, pigs and poultry is often practised by smallholders in low-income countries and is considered an important driver of influenza virus evolution. This study determined the prevalence and genetic characteristics of influenza A virus (IAV) in backyard pigs and poultry in Cambodia. A total of 751 animals were tested by matrix gene-based rRT-PCR, and influenza virus was detected in 1.5% of sampled pigs, 1.4% of chickens and 1.0% of ducks, but not in pigeons. Full-length genome sequencing confirmed triple reassortant H3N2 in all IAV-positive pigs and various low pathogenic avian influenza subtypes in poultry. Phylogenetic analysis of the swine influenza viruses revealed that these had haemagglutinin and neuraminidase genes originating from human H3N2 viruses previously isolated in South-East Asia. Phylogenetic analysis also revealed that several of the avian influenza subtypes detected were closely related to internal viral genes from highly pathogenic H5N1 and H9N2 formerly sequenced in the region. High sequence homology was likewise found with influenza A viruses circulating in pigs, poultry and wild birds in China and Vietnam, suggesting transboundary introduction and cocirculation of the various influenza subtypes. In conclusion, highly pathogenic subtypes of influenza virus seem rare in backyard poultry, but virus reassortment, involving potentially zoonotic and pandemic subtypes, appears to occur frequently in smallholder pigs and poultry. Increased targeted surveillance and monitoring of influenza circulation on smallholdings would further improve understanding of the transmission dynamics and evolution of influenza viruses in humans, pigs and poultry in the Mekong subregion and could contribute to limit the influenza burden.

Risk factors associated with Campylobacter detected by PCR in humans and animals in rural Cambodia.

Campylobacter are worldwide-occurring zoonotic bacteria, with the species Campylobacter jejuni and C. coli commonly associated with diarrhoea in children in low-income countries. In this cross-sectional study, the prevalence of C. jejuni and C. coli in human and livestock faecal samples was detected by PCR and zoonotic risk factors associated with human Campylobacter positivity were identified. In total 681 humans and 753 livestock (chickens, ducks, pigs, cattle) from 269 households were sampled. Children aged <16 years were more frequently Campylobacter positive (19%) than adults (8%) and multilevel logistic models revealed that human C. jejuni positivity was associated with the following household practices: home-slaughtering [odds ratio (OR) 2·4, P = 0·01], allowing animals access to sleeping and food preparation areas (OR 2·8, P = 0·02), and eating undercooked meat (OR 6·6, P = 0·05), while frequent consumption of beef was protective (OR 0·9, P = 0·05). Associations were stronger for home-slaughtering (OR 4·9, P = 0·004) with C. jejuni infection in children only. Campylobacter was highly prevalent in pigs (72%) and chickens (56%) and risk factors associated with human Campylobacter positivity were identified throughout the meat production chain. The findings underline the importance of studying source attributions throughout the production chain and the need for upgraded understanding of Campylobacter epidemiology in low-income countries.