Impaired pancreatic beta-cell function after a single dose of oral iron: A before-and-after (pre-post) study.

BACKGROUND: Although in vitro and animal studies have shown that iron loading in pancreatic beta cells impairs insulin secretion, no human studies have documented the acute effects of oral iron on beta-cell insulin secretory capacity. In the present study, we determined beta-cell insulin secretory capacity at baseline and after a single oral dose of iron (ferrous sulphate, 120 mg elemental iron) in healthy male individuals. METHODS: Fifteen healthy male volunteers underwent an oral glucose tolerance test (OGTT) to document baseline glucose tolerance and insulin secretion kinetics (baseline OGTT). One week later, the same subjects underwent a second OGTT, 2 h after an oral dose of ferrous sulphate (120 mg of elemental iron) (post-iron OGTT). Changes in disposition index, insulin secretion kinetics, glucose tolerance, insulin resistance, insulin clearance and iron-related parameters in serum were determined. RESULTS: Compared to baseline OGTT, the areas under the curve (AUC) for serum iron and transferrin saturation increased by 125% and 118%, respectively, in the post-iron OGTT. The disposition index decreased by 20% (p = 0.009) and the AUC for glucose concentrations increased by 5.7% (p < 0.001) during the post-iron OGTT. The insulin secretion rate was marginally lower during the first hour (-3.5%, p = 0.63), but became significantly higher during the second hour (22%, p = 0.005) of the post-iron OGTT. Insulin resistance and insulin clearance rate were not affected by iron intake. CONCLUSIONS: The decrease in disposition index and glucose tolerance observed after the oral dose of iron points to an acute iron-induced impairment in pancreatic beta-cell insulin secretory capacity.

Coping with autism during lockdown period of the COVID-19 pandemic: A cross-sectional survey.

BACKGROUND: The coronavirus disease 2019 (COVID-19) pandemic is thought to have greatly impacted families of individuals with autism spectrum disorder (ASD) due to lockdown, given lack of access to healthcare, therapy, and day-care centers. This survey was conducted to understand the magnitude of the impact of lockdown, and its effect on the health and behavior of individuals with ASD and their families. MATERIALS AND METHODS: We conducted an anonymous online survey, disseminated to families registered with our hospital and collaborating centers. The survey questionnaire collected information on sociodemographic details, details of the patient's and parents' behavior and health during the COVID-19 lockdown, and treatment details of the patient. RESULTS: A total of 153 families completed the survey. Of the 153, nearly half of the individuals with ASD had an inadequate understanding of lockdown, 54% had increased screen-time, while a third reported new-onset behavioral changes. About 40% received online therapies, of which 85% reported benefits. Of the 132 who answered the parent's section, 55% reported decreased interest and/or pleasure in doing daily activities and 43% felt depressed and/or hopeless. About 80% of families reported short-term positive changes such as improved speech, language skills, and participation in household chores. CONCLUSION: The COVID-19 pandemic has disrupted routines, triggered behavioral issues in individuals with ASD, and impacted the coping skills of both individuals and families, along with the mental health and well-being of the family. Valuable suggestions to improve therapy services and clinical care using technology have been uncovered and need to be explored.

Coexistence of mycobacterial infections - Mycobacterium tuberculosis and Mycobacterium leprae - in Sri Lanka: a case series.

BACKGROUND: Leprosy is one of the oldest mycobacterial infections and tuberculosis is the most common mycobacterial infection with a higher degree of infectivity than leprosy. Although both diseases are prevalent in clusters in developing countries, simultaneous occurrence of them in an individual is a rare entity, even in an endemic setting. CASE PRESENTATION: We describe six cases of tuberculosis and leprosy coinfection: a 57-year-old Sinhalese woman, a 47-year-old Tamil woman, a 72-year-old Tamil man, a 59-year-old Sinhalese man, a 54-year-old Sinhalese man, and a 50-year-old Sinhalese man. In this case series, five patients had lepromatous leprosy and the majority of patients were men. Three patients were detected to have tuberculosis at the outset of treatment of leprosy, while two developed tuberculosis later and one had extrapulmonary tuberculosis 5 years before the diagnosis of leprosy. The latter developed pulmonary tuberculosis as a reactivation while on treatment for leprosy. A majority of our patients with pulmonary tuberculosis had positive Mantoux test, high erythrocyte sedimentation rate, radiological evidence, and acid-fast bacilli in sputum. Human immunodeficiency virus and diabetes were detected in one patient. One patient had rifampicin-resistant tuberculosis, while she was on monthly rifampicin therapy for leprosy. CONCLUSION: An immunocompromised status, such as human immunodeficiency virus infection, diabetes, and immunosuppressive drugs, are risk factors for tuberculosis infection. The use of steroids in the treatment of leprosy may increase the susceptibility to develop tuberculosis. Development of rifampicin resistance secondary to monthly rifampicin in leprosy is a major concern in treating patients coinfected with tuberculosis. Despite the paucity of reports of coinfection, it is advisable to screen for tuberculosis in patients with leprosy, especially if there are respiratory or constitutional symptoms, high erythrocyte sedimentation rate, and abnormal chest X-ray. The fact is that positive Mantoux and QuantiFERON Gold tests and presence of acid-fast bacilli in sputum are misleading, chest X-ray evidence of active tuberculosis and positive tuberculosis cultures are important diagnostic clues for active tuberculosis infection in a patient with leprosy. This is important to avoid monthly rifampicin in patients with suspected coinfections, which may lead to development of drug resistance to tuberculosis treatment. Whether prolonged steroid therapy in leprosy is a risk factor for development of tuberculosis is still controversial.

Prevalence of Vitamin B12 Deficiency and Its Associated Risk Factors among Pregnant Women of Rural South India: A Community-based Cross-sectional Study.

INTRODUCTION: Vitamin B12 is essential for the normal functioning of the nervous system and for the formation of red blood cells. Vegetarian diet, low socioeconomic status, and social and religious reasons are known risk factors of its deficiency. Pregnant women, children, and the elderly are vulnerable groups. Indians have the highest prevalence, but the data among pregnant women in the rural setting is lacking. OBJECTIVES: The objective of this study is to assess the prevalence of Vitamin B12 deficiency and its associated factors among pregnant women of rural South India. MATERIALS AND METHODS: A cross-sectional study was conducted to recruit consecutive 120 multigravida women with ≤20 weeks of gestation, attending the mobile doctor run clinic of Kaniyambadi block, Vellore. A structured questionnaire was administered, and blood samples were collected. RESULTS: The prevalence of Vitamin B12 deficiency (<200 pgm/ml) and anemia (Hb ≤10.5 g/dL) was 55% and 17.5%, respectively. Only 11.7% were B12 deficient and anemic. Past history of abortion (odds ratio [OR] = 0.5), fatigue (OR = 0.4), and low B12 intake (OR = 2) was associated only in the bivariate analysis. First trimester (OR = 3.9) and obesity (OR = 9.6) were found to be independent risk factors of Vitamin B12 deficiency. CONCLUSION: Our study showed a high prevalence of Vitamin B12 deficiency in pregnancy in rural India. Some risk factors were identified. However, studies with a higher sample size will be beneficial to study the associated risk factors better.

Assessment of salivary endothelin-1 in patients with leukoplakia, submucous fibrosis, oral cancer and healthy individuals - a comparative study.

BACKGROUND: Oral cancer is one of the highly prevalent cancers worldwide being. According to data of GLOBOCAN 2018, the estimated incidence, mortality and 5-year survival rates due to lip, oral cavity and salivary gland cancer in world is (2.0%), (0.5%) and (0.3%) respectively. (Bray, Ferlay and Soerjomataram, 2018). Endothelin-1 (ET-1) is a 21-amino acid peptide; its receptors have been implicated in the growth and progression of both primary and metastatic neoplasms throughout the human body. Studies have shown that ET-1 is expressed in tissue, serum and other body fluids. AIM: To estimate the levels of salivary endothelin-1 in Oral potentially malignant disorders (oral leukoplakia and submucous fibrosis) and oral squamous cell carcinoma. MATERIALS AND METHODS: The study population included 60 subjects and were divided into 4 groups. All patients included in the study are clinically and histopathological diagnosed cases of oral leukoplakia, submucous fibrosis and oral cancer and assessed for salivary ET-1 levels using human ELISA kit. Significant differences between the groups were determined using one-way analysis of variance, LSD and Post HOC, unpaired t test, biserial and spearson's correlation. RESULTS: The mean levels of salivary Endothelin-1 level in study groups were: 82.78 ± 5.9 pg/mL (OSCC), 65.02 ± 1.8 pg/mL (SMF), 57.76 ± 4.1 pg/mL (LEUKOPLAKIA), 29.72 ± 14.1 pg/mL (CONTROLS). The mean Salivary ET-1 levels among these four groups was compared and the difference was statistically significant (P < 0.001). We also found a significant difference in the means of ET-1 levels among the clinical and histopathological staging of the study groups. CONCLUSION: Our results demonstrate potential utility of salivary analysis for ET-1 levels to monitor patients at risk for OSCC. Although provides the basis for a larger prospective study to determine the critical levels of salivary ET-1 necessary to diagnose and monitor OPMD's and its potential to undergo malignant transformation.

Urine and serum fetuin-A levels in patients with urolithiasis.

INTRODUCTION: Fetuin-A is a glycoprotein secreted by liver and has been shown to inhibit extraosseous mineralization. Urolithiasis may be a manifestation in the urinary tract due to fetuin deficiency in urine. The objective of this study was to compare the 24-h urine and serum fetuin-A levels of patients with and without urolithiasis. METHODS: Serum and 24-h urine fetuin-A levels were measured in 41 patients with bilateral, multiple, or recurrent urinary tract calculi (Group A) and 41 matched controls with no calculi (Group B). Fetuin levels were measured by enzyme linked immunosorbent assay. Serum and urine fetuin-A levels in the two groups were compared. RESULTS: The median (range) 24-h urine fetuin-A value in Group A was 11.9 (1.12-221) mg/day and in Group B was 37.7 (1.28-125) mg/day. This difference was statistically significant (Mann-Whitney test, P = 0.0169). The median (range) serum fetuin-A in Group A was 0.67 (0.05-2.68) g/L and in Group B was 0.99 (0.01-5.5) g/L. The difference between serum values in the two arms was not statistically significant (Mann-Whitney test, P = 0.1817). However, the serum creatinine-adjusted mean log serum fetuin and urine fetuin were significantly different in the two arms (P = 0.003). The mean ± standard deviation (range) serum creatinine in Group A was 0.98 ± 0.25 (0.56-1.58) mg% and in Group B was 0.83 ± 0.16 (0.58-1.18) mg% (two sample t-test, P = 0.0031). CONCLUSIONS: Patients with urolithiasis have lower urine fetuin-A and creatinine-adjusted serum fetuin-A levels.

Molecular Methods and Bioinformatic Tools for Adjuvant Characterization by High-Throughput Sequencing.

Adjuvants in vaccine formulations are designed to enhance immune responses against a target antigen or pathogen. The ability of these vaccines to induce activation and differentiation of mature naïve B cells to produce pathogen-specific antibodies (immunoglobulins; Ig) helps guarantee long-lived humoral immunity. This process involves clonal expansion of antigen-specific B cells, genomic rearrangement of Ig heavy (IgH) and light (IgL) loci, somatic hypermutation (SHM), and clonal selection for affinity-matured antibody, resulting in a vast but directed repertoire of B cells expressing highly specific antibody proteins. High-throughput sequencing of the IgH and IgL complementary determining regions (CDRs) derived from various B cell populations provides an unprecedented way to observe dynamic responses of the humoral immune repertoire in response to vaccination. However, applying high-throughput sequencing (HTS) methodologies to multi-armed in vivo experiments requires careful coordination of sample preparation with downstream bioinformatics, particularly with regard to issues of quantitation, sequence fidelity, bar-coding, and multiplexing strategies. Here, we overview strategies of high-throughput sequencing and analysis of the adaptive immune complex loci applied to multi-armed, multiplexed experiments.

From mouse to man: safety, immunogenicity and efficacy of a candidate leishmaniasis vaccine LEISH-F3+GLA-SE.

Key antigens of Leishmania species identified in the context of host responses in Leishmania-exposed individuals from disease-endemic areas were prioritized for the development of a subunit vaccine against visceral leishmaniasis (VL), the most deadly form of leishmaniasis. Two Leishmania proteins-nucleoside hydrolase and a sterol 24-c-methyltransferase, each of which are protective in animal models of VL when properly adjuvanted- were produced as a single recombinant fusion protein NS (LEISH-F3) for ease of antigen production and broad coverage of a heterogeneous major histocompatibility complex population. When formulated with glucopyranosyl lipid A-stable oil-in-water nanoemulsion (GLA-SE), a Toll-like receptor 4 TH1 (T helper 1) promoting nanoemulsion adjuvant, the LEISH-F3 polyprotein induced potent protection against both L. donovani and L. infantum in mice, measured as significant reductions in liver parasite burdens. A robust immune response to each component of the vaccine with polyfunctional CD4 TH1 cell responses characterized by production of antigen-specific interferon-γ, tumor necrosis factor and interleukin-2 (IL-2), and low levels of IL-5 and IL-10 was induced in immunized mice. We also demonstrate that CD4 T cells, but not CD8 T cells, are sufficient for protection against L. donovani infection in immunized mice. Based on the sum of preclinical data, we prepared GMP materials and performed a phase 1 clinical study with LEISH-F3+GLA-SE in healthy, uninfected adults in the United States. The vaccine candidate was shown to be safe and induced a strong antigen-specific immune response, as evidenced by cytokine and immunoglobulin subclass data. These data provide a strong rationale for additional trials in Leishmania-endemic countries in populations vulnerable to VL.

Issues related to obtaining intelligence quotient-matched controls in autism research.

BACKGROUND: Intelligence Quotient (IQ) is considered to be an index of global cognitive functioning and has traditionally been used as a fulcral measure in case-control studies in neuro-developmental disorders such as autism. AIM: The aim is to highlight the issues of "matching for IQ" with controls in autism research. MATERIALS AND METHODS: Percentile scores on the Coloured Progressive Matrices of 20 children with autism in the age range of 5 to 12 years have been graphically compared with 21 age matched typically developing children. RESULTS AND CONCLUSIONS: The percentile scores of the so-called high functioning children with autism from special schools were well below that of typically developing children. There are many challenges when using IQ in case-control studies of autism. Alternative approaches need to be considered.

CCR6-dependent positioning of memory B cells is essential for their ability to mount a recall response to antigen.

Chemokine-dependent localization of specific B cell subsets within the immune microarchitecture is essential to ensure successful cognate interactions. Although cognate interactions between T cells and memory B cells (B(mem)) are essential for the secondary humoral immune responses, the chemokine response patterns of B(mem) cells are largely unknown. In contrast to naive B cells, this study shows that Ag-specific B(mem) cells have heightened expression of CCR6 and a selective chemotactic response to the CCR6 ligand, CCL20. Although CCR6 appears be nonessential for the initial clonal expansion and maintenance of B(mem), CCR6 is essential for the ability of B(mem) to respond to a recall response to their cognate Ag. This dependency was deemed intrinsic by studies in CCR6-deficient mice and in bone marrow chimeric mice where CCR6 deficiency was limited to the B cell lineage. Finally, the mis-positioning of CCR6-deficient B(mem) was revealed by immunohistological analysis with an altered distribution of CCR6-deficient B(mem) from the marginal and perifollicular to the follicular/germinal center area.

Regulatory actions of Toll-like receptor 2 (TLR2) and TLR4 in Leishmania donovani infection in the liver.

In livers of susceptible but self-curing C57BL/6 mice, intracellular Leishmania donovani infection enhanced Toll-like receptor 4 (TLR4) and TLR2 gene expression. In the liver, infected TLR4(-/-) mice showed reduced gamma interferon (IFN-γ), tumor necrosis factor (TNF), and inducible nitric oxide synthase (iNOS) mRNA expression, higher-level and slowly resolving infection, delayed granuloma formation, and little response to low-dose chemotherapy; in serum, the ratio of IFN-γ to interleukin 10 (IL-10) activity was decreased by 50%. In contrast, in TLR2(-/-) mice, control of liver infection, parasite killing, and granuloma assembly were accelerated and chemotherapy's efficacy enhanced. In livers of infected TLR2(-/-) mice, mRNA expression was not increased for inflammatory cytokines or iNOS or decreased for IL-10; however, the serum IFN-γ/IL-10 ratio was increased 6.5-fold and minimal responses to IL-10 receptor blockade suggested downregulated IL-10. In established infection in wild-type mice, blockading TLR2 induced parasite killing and triggering TLR4 strengthened resistance and promoted chemotherapy's effect. Thus, in experimental L. donovani infection in the liver, TLR4 signaling upregulates and TLR2 signaling downregulates macrophage antileishmanial activity, making both receptors potential therapeutic targets in visceral leishmaniasis for engagement (TLR4) or blockade (TLR2).

Development and pre-clinical assessment of a 73 kD chimeric fusion protein as a defined sub-unit vaccine for leprosy.

Despite the advances toward the elimination of leprosy through widespread provision of multi-drug therapy to registered patients over the last 2 decades, new case detection rates have stabilized and leprosy remains endemic in a number of localized regions. A vaccine could overcome the inherent limitations of the drug treatment program by providing protection in individuals who are not already harboring the Mycobacterium leprae bacilli at the time of administration and effectively interrupt the transmission cycle over a wider timespan. In this report we present data validating the production of 73f, a chimeric fusion protein incorporating the M. leprae antigens ML2028, ML2346 and ML2044. The 73f protein was recognized by IgG in multibacillary (MB) leprosy patient sera and stimulated IFNγ production within whole blood assays of paucibacillary (PB) leprosy patient and healthy household contacts of MB patients (HHC). When formulated with a TLR4L-containing adjuvant (GLA-SE), 73f stimulated a strong and pluripotent Th1 response that inhibited M. leprae-induced inflammation in mice. We are using these data to develop new vaccine initiatives for the continued and long-term control of leprosy.

Adjuvants for Leishmania vaccines: from models to clinical application.

Two million new cases of leishmaniasis occur every year, with the cutaneous leishmaniasis (CL) presentation accounting for approximately two-thirds of all cases. Despite the high incidence rates and geographic expansion of the disease, CL remains a neglected tropical disease without effective intervention strategies. Efforts to address this deficit have given rise to the experimental murine model of CL. By virtue of its simplicity and pliability, the CL model has been used to provide substantial information regarding cellular immunity, as well as in the discovery and evaluation of various vaccine adjuvants. The CL model has facilitated in vivo studies of the mechanism of action of many adjuvants, including the TLR4 agonist monophosphoryl lipid A, the TLR7/8 agonist imiquimod, the TLR9 agonist CpG, adenoviral vectors, and the immunostimulatory complexes. Together, these studies have helped to unveil the requirement for certain types of immune responses at specific stages of CL disease and provide a basis to aid the design of effective second-generation vaccines for human CL. This review focuses on adjuvants that have been tested in experimental CL, outlining how they have helped advance our understanding of the disease and ultimately, how they have performed when applied within clinical trials against human CL.

The development and clinical evaluation of second-generation leishmaniasis vaccines.

Infection with Leishmania parasites results in a range of clinical manifestations and outcomes. Control of Leishmania parasite transmission is extremely difficult due to the large number of vectors and potential reservoirs, and none of the current treatments are ideal. Vaccination could be an effective strategy to provide sustained control. In this review, the current global situation with regard to leishmaniasis, the immunology of Leishmania infection and various efforts to identify second generation vaccine candidates are briefly discussed. The variety of clinical trials conducted using the only current second generation vaccine approved for clinical use, LEISH-F1+MPL-SE, are described. Given that epidemiological evidence suggests that reducing the canine reservoir also positively impacts human incidence, efforts at providing a vaccine for leishmaniasis in dogs are highlighted. Finally, potential refinements and surrogate markers that could expedite the introduction of a vaccine that can limit the severity and incidence of leishmaniasis are discussed.

Targeting TLRs expands the antibody repertoire in response to a malaria vaccine.

Vaccination with an isolated antigen is frequently not sufficient to elicit a protective immune response. The addition of adjuvants to the antigen can increase the magnitude and breadth of the response generated, but quantification of this increase as a function of adjuvant has been intractable. We have directly determined the variation of the immunoglobulin G variable-chain repertoire of an entire organism as a function of vaccination. Using the well-established Plasmodium vivax antigen, PvRII, and massively parallel sequencing, we showed that the use of a Toll-like receptor (TLR) agonist in the vaccine formulation increased the diversity of the variable region sequences in comparison to the use of an oil-in-water emulsion adjuvant alone. Moreover, increased variable domain diversity in response to the use of TLR agonist-based adjuvants correlated with improved antigen neutralization. The use of TLR agonists also broadened the range of polymorphic variants against which these antibodies could be effective. In addition, a peptide microarray demonstrated that inclusion of adjuvants changed the profile of linear epitopes from PvRII that were recognized by serum from immunized animals. The results of these studies have broad implications for vaccine design--they may enable tailored adjuvants that elicit the broad spectrum of antibodies required to neutralize drifted and polymorphic pathogen strains as well as provide a method for rapid determination of correlates of adjuvant-induced humoral immunity.

KSAC, the first defined polyprotein vaccine candidate for visceral leishmaniasis.

A subunit vaccine using a defined antigen(s) may be one effective solution for controlling leishmaniasis. Because of genetic diversity in target populations, including both dogs and humans, a multiple-antigen vaccine will likely be essential. However, the cost of a vaccine to be used in developing countries must be considered. We describe herein a multiantigen vaccine candidate comprised of antigens known to be protective in animal models, including dogs, and to be recognized by humans immune to visceral leishmaniasis. The polyprotein (KSAC) formulated with monophosphoryl lipid A, a widely used adjuvant in human vaccines, was found to be immunogenic and capable of inducing protection against Leishmania infantum, responsible for human and canine visceral leishmaniasis, and against L. major, responsible for cutaneous leishmaniasis. The results demonstrate the feasibility of producing a practical, cost-effective leishmaniasis vaccine capable of protecting both humans and dogs against multiple Leishmania species.

A synthetic TLR4 agonist formulated in an emulsion enhances humoral and Type 1 cellular immune responses against GMZ2--a GLURP-MSP3 fusion protein malaria vaccine candidate.

GMZ2 adjuvanted by aluminum hydroxide is a candidate malaria vaccine that has successfully passed phase 1 clinical testing in adult German and Gabonese volunteers and Gabonese children under five. Here we report a preclinical study screening a series of adjuvant vehicles and Toll-like receptor (TLR) agonists in CB6F1 mice to identify an improved formulation of GMZ2 suitable for further human clinical studies. GMZ2 formulated in an oil-in-water emulsion plus the synthetic TLR4 agonist GLA elicits the highest (a) vaccine-specific IgG2a and total IgG titers, (b) parasite-specific IFA titers, (c) levels of Type 1 cytokine responses (IFN-γ), and (d) number of long-lived-plasma cells (LLPC) secreting antibodies against both the GMZ2 fusion and its two components. Thus, GLA helps to elicit a vaccine-specific Type 1 antibody profile together with high levels of LLPC, both of which are thought to be essential for the development of long-term protective immunity against clinical malaria.

A synthetic adjuvant to enhance and expand immune responses to influenza vaccines.

Safe, effective adjuvants that enhance vaccine potency, including induction of neutralizing Abs against a broad range of variant strains, is an important strategy for the development of seasonal influenza vaccines which can provide optimal protection, even during seasons when available vaccines are not well matched to circulating viruses. We investigated the safety and ability of Glucopyranosyl Lipid Adjuvant-Stable Emulsion (GLA-SE), a synthetic Toll-like receptor (TLR)4 agonist formulation, to adjuvant Fluzone® in mice and non-human primates. The GLA-SE adjuvanted Fluzone vaccine caused no adverse reactions, increased the induction of T helper type 1 (T(H)1)-biased cytokines such as IFNγ, TNF and IL-2, and broadened serological responses against drifted A/H1N1 and A/H3N2 influenza variants. These results suggest that synthetic TLR4 adjuvants can enhance the magnitude and quality of protective immunity induced by influenza vaccines.