Lipid-induced polymorphic amyloid fibril formation by α-synuclein.

Many proteins that self-assemble into amyloid and amyloid-like fibers can adopt diverse polymorphic forms. These forms have been observed both in vitro and in vivo and can arise through variations in the steric-zipper interactions between ß-sheets, variations in the arrangements between protofilaments, and differences in the number of protofilaments that make up a given fiber class. Different polymorphs arising from the same precursor molecule not only exhibit different levels of toxicity, but importantly can contribute to different disease conditions. However, the factors which contribute to formation of polymorphic forms of amyloid fibrils are not known. In this work, we show that in the presence of 1,2-dimyristoyl-sn-glycero-3-phospho-L-serine, a highly abundant lipid in the plasma membrane of neurons, the aggregation of α-synuclein is markedly accelerated and yields a diversity of polymorphic forms under identical experimental conditions. This morphological diversity includes thin and curly fibrils, helical ribbons, twisted ribbons, nanotubes, and flat sheets. Furthermore, the amyloid fibrils formed incorporate lipids into their structures, which corroborates the previous report of the presence of α-synuclein fibrils with high lipid content in Lewy bodies. Thus, the present study demonstrates that an interface, such as that provided by a lipid membrane, can not only modulate the kinetics of α-synuclein amyloid aggregation but also plays an important role in the formation of morphological variants by incorporating lipid molecules in the process of amyloid fibril formation.

Tracheostomy in Pediatric Intensive Care Unit-A Two Decades of Experience.

AIM AND OBJECTIVE: To study the profile, indications, related complications, and predictors of decannulation and mortality in patients who underwent tracheostomy in the pediatric intensive care unit (PICU). MATERIALS AND METHODS: Retrospective analysis of prospectively collected data of tracheostomies was done on patients admitted at PICU. Demographics, primary diagnosis, indication of tracheostomy, and durations of endotracheal intubation, mechanical ventilation, and tracheostomy cannulation were recorded. The indication was recorded in one of the four categories-upper airway obstruction (UAO), central neurological impairment (CNI), prolonged mechanical ventilation, and peripheral neuromuscular disorders). RESULTS: Two hundred ninety cases were analyzed. UAO (42%) and CNI (48.2%) were main indications in the halves of the study period, respectively. Decannulation was successful in 188 (64.8%) patients. Seventy-seven percentage UAO patients were decannulated successfully [OR (odds ratio); 95% CI (confidence interval), 2.647; 1.182-5.924, p = 0.018]. Age <1 year (0.378; 0.187-0.764; p = 0.007), nontraumatic, noninfectious central neurological diseases (0.398; 0.186-0.855; p = 0.018), and malignancy (0.078; 0.021-0.298; p <0.001), durations of posttracheostomy ventilation (0.937; 0.893-0.983; p = 0.008), and stay in the PICU (0.989; 0.979-0.999; p = 0.029) were predictors of unsuccessful decannulation. There were 91 (31.4%) deaths. Age <1 year (2.39 (1.13-5.05; p = 0.02), malignancy (17.55; 4.10-75.11; p <0.001), durations of posttracheostomy ventilation (1.06; 1.006-1.10; p = 0.028), and hospital stay (1.007; 1.0-1.013; p = 0.043) were independent predictors of mortality. Indication of UAO favored survivor (0.24; 0.09-0.57; p <0.001). CONCLUSION: The indications for tracheostomy in children had changed over the years. Infancy, primary diagnosis, length of posttracheostomy ventilation, and stay in the PICU and hospital were independent predictors of decannulation and mortality. WHAT THIS ADDS: Similar to developed countries, the age at the time of tracheostomy and indication are changing. Inability to decannulate and mortality were associated with the age of a child at the time of tracheostomy, indication, medical diagnosis, and duration of postprocedure mechanical ventilation and stay in the hospital. HOW TO CITE THIS ARTICLE: Sachdev A, Chaudhari ND, Singh BP, Sharma N, Gupta D, Gupta N, et al. Tracheostomy in Pediatric Intensive Care Unit-A Two Decades of Experience. Indian J Crit Care Med 2021;25(7):803-811.

Giant pressure sensitivity in piezo/ferro-electric ceramics.

We report the fabrication of single-phase polycrystalline Pb0.85Bi0.10(Zr0.52Ti0.48)O3 (PBiZT) ceramic which shows large polarization, i.e., ∼40 µC cm-2 and piezoelectric coefficients ∼130 pC N-1 and giant linear change in capacitive reactance and dielectric properties with increasing and decreasing pressure in the range of 1 kHz to 5 MHz. Nearly 70% change in dielectric constant and 56% change in capacitive reactance were obtained in the pressure range of 20-200 MPa, which makes it suitable for applications as a capacitive pressure sensor/gauge. The sensitivity of the device is calculated as 0.66 MPa-1 and 18.2 MPa-1 at 1 MHz and 5 MHz, respectively, which is the highest ever reported value so far for any bulk polycrystalline ceramic. The compressive stress of the device was tested according to the standard test method as a function of linear and volumetric strain, which yields the Young's modulus, Bulk modulus, and Poisson's ratio of the device. These values were further utilized to calculate actual stress in the sample and energy density using ANSYS software, which indicates at least four orders smaller pressure in the sample compared to the applied pressure.

Improving Production of Tacrolimus In Streptomyces Tacrolimicus (ATCC 55098) Through Development of Novel Mutant by Dual Mutagenesis

ABSTRACT Tacrolimus is a polyketide macrolide produced by Streptomyces species which is widely used as anti-fibrotic agent and potent immunosuppressant. In this article dual mutagenesis approach using mutagens (NTG+EMS+UV) was used to develop a mutant strain of Streptomyces tacrolimicus (ATCC 55098) for higher tacrolimus production and this strain showed higher tacrolimus production at 82.5 mg/l. Interestingly; addition of L-Lysine (0.2 g/l) into the production medium further enhanced the tacrolimus production to ~102 mg/l at 7-L fed-batch bioreactor. To the best of our knowledge this is the first report mentioning efficient strain development for higher production of tacrolimus using dual mutagenesis. The obtained data presents an impressive model for higher production of tacrolimus and enhanced our understanding regarding improvement in production capacity of tacrolimus in Streptomyces tacrolimicus.

Proceedings of the 2014 National Toxicology Program Satellite Symposium.

The 2014 annual National Toxicology Program (NTP) Satellite Symposium, entitled "Pathology Potpourri" was held in Washington, D.C., in advance of the Society of Toxicologic Pathology's 33rd annual meeting. The goal of this annual NTP Symposium is to present current diagnostic pathology or nomenclature issues to the toxicologic pathology community. This article presents summaries of the speakers' presentations, including diagnostic or nomenclature issues that were presented, along with select images that were used for audience voting and discussion. Some lesions and topics covered during the symposium included a pulmonary mucinous adenocarcinoma in a male B6C3F1 mouse; plexiform vasculopathy in Wistar Han (Crl:WI[Han]) rats; staging of the estrous cycle in rats and mice; peri-islet fibrosis, hemorrhage, lobular atrophy and inflammation in male Sprague-Dawley (SD) rats; retinal dysplasia in Crl:WI[Han] rats and B6C3F1 mice; multicentric lymphoma with intravascular microemboli and tumor lysis syndrome, and 2 cases of myopathy and vascular anomaly in Tg.rasH2 mice; benign thymomas in Crl:WI[Han] rats; angiomatous lesions in the mesenteric lymph nodes of Crl:WI[Han] rats; an unusual foveal lesion in a cynomolgous monkey; and finally a series of nomenclatures challenges from the endocrine International Harmonization of Nomenclature and Diagnostic Criteria (INHAND) Organ Working Group (OWG).

Mice deficient in glycerol-3-phosphate acyltransferase-1 have a reduced susceptibility to liver cancer.

The risk of hepatocellular carcinoma increases with the persistence of non-alcoholic fatty liver disease. Triacylglycerol synthesis is initiated by glycerol-3-phosphate acyltransferase (GPAT). Of four isoforms, GPAT1 contributes 30-50% of total liver GPAT activity, and we hypothesized that it might influence liver susceptibility to tumorigenesis. C57Bl/6 mice deficient in GPAT1 were backcrossed 6 times to C3H mice. After exposure to the carcinogen diethylnitrosamine (DEN) and the tumor promoter phenobarbital, male Gpat1⁻/⁻ mice, compared with controls (Gpat1⁺/⁺), had 93% fewer macroscopically visible nodules per liver at 21 weeks of age and 39% fewer at 34 weeks of age. Microscopically, control mice had increased numbers of foci of altered hepatocytes, particularly the basophilic subtype, as well as more, and malignant, liver neoplasms than did the Gpat1⁻/⁻ mice. At 21 weeks of age, 50% (4/8) of control mice (50%) had hepatocellular adenomas with an average multiplicity (tumors per tumor-bearing-animal) of 4.3, while none occurred in 8 Gpat1⁻/⁻ mice. At 34 weeks of age, all 15 control mice (100%) had hepatocellular adenomas with an average multiplicity of 5.2 compared to an incidence of 93% in Gpat1⁻/⁻ mice and multiplicity of 3.1. HCCs were observed in 13% of control mice and in only 6% of Gpat1⁻/⁻ mice. These data show that alterations in the formation of complex lipids catalyzed by Gpat1 reduce susceptibility to DEN-induced liver tumorigenesis.

Current and future use of genomics data in toxicology: opportunities and challenges for regulatory applications.

Toxicogenomics is the application of toxicology, genetics, molecular biology and environmental health to describe the response of organisms to environmental stimuli. The field of toxicogenomics has developed over the past 15 years mainly due to advances in toxicology, molecular genetics and cell biology. Its prospective use to resolve crucial data gaps and data inconsistencies could improve risk assessment by providing additional data to increase the understanding of mechanisms and modes of action (MOA) and enhance the reliability of dose-response extrapolation. Thus, toxicogenomics holds promise for advancing the scientific basis of risk assessments. However, one of the current issues is how genomic/transcriptional data is being used to further describe a MOA for oncogenicity and, in turn, its potential uses in cancer risk assessment. This commentary identifies how toxicogenomics could be used on a case by case basis to add information to a MOA addressing both the opportunities and challenges this technology holds. In addition, some pitfalls to avoid in the generation and interpretation of toxicogenomic data and validation issues that need to be addressed before toxicogenomics can be used in the risk assessment process and regulatory decisions are discussed.

Choline attenuates immune inflammation and suppresses oxidative stress in patients with asthma.

Asthma is a chronic immune inflammatory disease characterized by variable airflow obstruction and increased bronchial hyperreactivity (BHR). Therapeutic interventions reduce airway inflammation and relieve symptoms but associated with potential side effects that limit their usefulness. The present study was undertaken to assess the effect of choline on immune inflammation and BHR in asthma subjects. The patients of asthma (n=76) were recruited and treated with choline supplement (1500 mg twice) or standard pharmacotherapy for 6 months in two groups. The patients were evaluated by clinical, immunologic and biochemical parameters. The treatment with choline showed significant reduction in symptom/drug score and improvement in PC(20) FEV1 compared to baseline or standard pharmacotherapy (p<0.01). Choline therapy significantly reduced IL-4, IL-5 and TNF-alpha level as compared to baseline or standard pharmacotherapy after 6 months (p<0.01). Blood eosinophil count and total IgE levels were reduced in both the treatment groups. Cysteinyl leukotriene and leukotriene B4 were suppressed significantly by choline treatment (p<0.01). This was accompanied by decreased 8-isoprostanes, a biomarker for oxidative stress after choline treatment (p<0.01). Choline therapy modulates immune inflammation and suppresses oxidative stress in asthma patients. It can be used as an adjunct therapy for asthma patients.

Clinico-immunologic study on immunotherapy with mixed and single insect allergens.

BACKGROUND: Immunotherapy (IT) is practiced mainly with mixed and single allergen vaccines. But studies are rare with mixed allergen preparations. OBJECTIVE: The objective of this study is to study mix and single insect allergen IT in patients of allergic rhinitis and asthma. METHODS: We performed a double-blind placebo-controlled trial of mix and single allergen IT for 1 year in 99 patients of asthma or rhinitis or both. There were two groups, (1) active allergen IT (n = 61) with three subgroups single insect extract (cockroach, housefly, or mosquito) and mix allergen IT (two or three insect extracts) and (2) placebo (n = 38). Clinical (skin reactivity, airway reactivity, and symptom score) and immunological (IgE/IgG4 and IgG1/IgG4 ratio) parameters were assessed at baseline and after 1 year of IT. RESULTS: Eighty-five patients completed 1 year of IT. The active allergen IT group patients showed a significant improvement compared to baseline values (p < 0.05) and placebo group patients (p < 0.05) with regard to symptom scores, FEV1 values, and immunological parameters (IgG4). No significant difference was found between mixed and single IT group patients for changes in clinical and immunological parameters. Positive correlation was observed between increase in IgG4 and clinical improvement. The changes in above parameters in placebo group were nonsignificant after 1 year of treatment. CONCLUSION: IT with two to three mix extract from the same allergen group is effective for insect hypersensitivity.

Bioinformatics and immunologic investigation on B and T cell epitopes of Cur l 3, a major allergen of Curvularia lunata.

The knowledge on epitopes of proteins can help in devising new therapeutic modalities for allergic disorders. In the present study, five B (P1-P5) and five T cell (P6-P10) epitopes were predicted in silico based on sequence homology model of Cur l 3, a major allergen of Curvularia lunata. Peptides (epitopes) were synthesized and assessed for biological activity by ELISA, competitive ELISA, lymphoproliferation and cytokine profiling using Curvularia allergic patients' sera. B cell peptides showed higher IgE binding by ELISA than T cell epitopes except P6. Peptides P1-P6 achieved EC(50) at 100 ng, whereas P7-P10 required 10 mug in inhibition assays. Peripheral blood mononuclear cells from Curvularia allergic patients (n = 20) showed higher lymphoproliferation for T cell epitopes than B cell epitopes except P6 confirming the properties of B and T cell prediction. The supernatant from these patients show highest interleukin-4 release on stimulation with P6 followed by B cell peptides. P4 and P6 together identified 35/37 of Curvularia positive patients by skin tests. In summary, experimental analysis confirmed in silico predicted epitopes containing important antigenic regions of Cur l 3. P6, a predicted T cell epitope, showed the presence of a cryptic B cell epitope. Peptides P4 and P6 have potential for clinical application. The approach used here is relevant and may be used to delineate epitopes of other proteins.

Effect of smoking on atopic predisposition and sensitisation to allergens.

BACKGROUND: Previous studies elsewhere have shown higher serum immunoglobulin E (IgE) levels in smokers and an association between smoking and sensitisation to allergens. Such information is not available for Indian population. Hence, the present study was carried out to evaluate the effect of smoking on atopic predisposition and sensitisation to aeroallergens. METHODS: A total of 70 subjects were included in the present study comprising of 25 smokers, 22 reformed smokers and 23 non-smokers. Absolute eosinophil count (AEC), serum total IgE levels and skin prick test (SPT) against common aeroallergens were performed in all the subjects along with breath carbon monoxide (CO) levels and pulmonary function tests. RESULTS: Smokers showed significantly higher serum total IgE levels (328.80 +/- 161.82 IU/mL) as compared to reformed smokers 177.27 +/- 86.47 IU/mL) and non-smokers (29.56 +/- 9.75 IU/mL). A number of subjects among smokers and reformed smokers elicited positive SPT reactions to various allergen extracts. Non-smokers did not show any significant positive skin reaction. The AECs were slightly higher in smokers (350 +/- 1145.61/mm3) as compared to reformed smokers (305 +/- 146.33/ mm3). Breath CO was considerably higher (greater than three times) in smokers than reformed smokers. However, reformed smokers showed greater airways obstruction than smokers. The former also had higher Brinkman index (646.81 +/- 392.32) as compared to the latter (448.36 +/- 279.86). CONCLUSIONS: Smokers had significantly higher IgE serum levels than reformed smokers and non-smokers. Smoking seems to induce an atopic orientation and allergen sensitisation in individuals.

Rostral cerebellar arterial infarct in two cats.

A 10-year-old female neutered domestic shorthair (DSH) cat and a 6-year-old female neutered Siamese cat were presented following a peracute onset of decerebellate rigidity and a cerebellar vestibular syndrome, respectively. In both cats, physical examination and routine blood tests were unremarkable, as was routine analysis of cerebrospinal fluid obtained from the DSH cat. Based on the magnetic resonance imaging (MRI) features - focal wedge-shaped lesion in the cerebellum characterised by hyperintensity in T2-weighted, T2( *)-gradient echo and fluid attenuated inversion recovery (FLAIR) images - a presumptive diagnosis of cerebellar infarct was made in both cases. In the DSH cat, the post-mortem examination confirmed the diagnosis of cerebellar infarct and additionally found acute renal infarcts and a pulmonary neoplasia. In the Siamese cat, ultrasonographic evaluation of the heart revealed a probable low-grade chronic valvular endocarditis which was thought to be a potential source of thromboembolism. This paper describes the first two cases - one confirmed and the other suspected - of cerebellar infarct in the cat. The in vivo potential diagnostic value of the MRI study is highlighted. Cerebellar infarcts should be included in the differential diagnosis of cat with a peracute onset of cerebellar signs regardless of the severity of neurological deficits.

Intrarenal pelvic nephroblastoma in a meerkat (Suricata suricatta).

Nephroblastoma is the most common primary renal tumor in children and has also been reported in domestic and nondomestic animal species. Intrapelvic renal nephroblastoma is a rare variant of this tumor type in human patients. Postmortem examination of a captive meerkat (Suricata suricatta), which was found dead, revealed enlargement of the pelvis of the left kidney by a tumor mass. Gross, histological, and immunohistochemical findings were consistent with a diagnosis of triphasic intrapelvic renal nephroblastoma. This is the first reported spontaneous case of intrapelvic renal nephroblastoma in a nonhuman species.