Prevalence of celiac disease in nutritional anemia at a tertiary care center

Background

While anemia occurs in 80 % to 90 % of patients with celiac disease (CD), it may be the sole manifestation of CD. The prevalence of CD in Indian patients with nutritional anemia is not known.

Patients and Methods

Adolescent and adult patients presenting with nutritional anemia were prospectively screened for CD using IgA anti-tissue transglutaminase antibody (anti-tTG Ab) followed, if positive, by upper gastrointestinal endoscopy and duodenal biopsy.

Results

Ninety-six patients [mean?±?SD age 32.1?±?13.1 years and median duration of anemia 11 months (range 1 to 144 months)] were screened. Of these patients, 80 had iron deficiency anemia, 11 had megaloblastic anemia, and 5 had dimorphic anemia. Seventy-three patients were on hematinics and 36.4 % had received blood transfusions. Nineteen had a history of chronic diarrhea and the mean?±?SD duration of diarrhea in them was 9.7?±?35.8 months. IgA anti-tTG Ab was positive in 13 patients, of whom 12 agreed to undergo duodenal biopsy. Ten patients had villous atrophy (Marsh grade 3a in three, 3b in one, and 3c in six) and two did not. Thus, 10 patients with nutritional anemia (iron deficiency 9, vitamin B₁₂ deficiency 1) were diagnosed to have CD. On multivariate logistic regression, age, duration of symptoms, and presence of diarrhea were found to be the predictors of CD. All the patients with CD were put on gluten-free diet and with iron and vitamin supplementations and showed a significant improvement in hemoglobin concentration.

Conclusions

CD screening should be included in the work up of otherwise unexplained nutritional anemia.     

Influenza hemagglutinin stem-fragment immunogen elicits broadly neutralizing antibodies and confers heterologous protection

Influenza hemagglutinin (HA) is the primary target of the humoral response during infection/vaccination. Current influenza vaccines typically fail to elicit/boost broadly neutralizing antibodies (bnAbs), thereby limiting their efficacy. Although several bnAbs bind to the conserved stem domain of HA, focusing the immune response to this conserved stem in the presence of the immunodominant, variable head domain of HA is challenging. We report the design of a thermotolerant, disulfide-free, and trimeric HA stem-fragment immunogen which mimics the native, prefusion conformation of HA and binds conformation specific bnAbs with high affinity. The immunogen elicited bnAbs that neutralized highly divergent group 1 (H1 and H5 subtypes) and 2 (H3 subtype) influenza virus strains in vitro. Stem immunogens designed from unmatched, highly drifted influenza strains conferred robust protection against a lethal heterologous A/Puerto Rico/8/34 virus challenge in vivo. Soluble, bacterial expression of such designed immunogens allows for rapid scale-up during pandemic outbreaks.Seasonal influenza outbreaks across the globe cause an estimated 250,000–500,000 deaths annually (1). Current influenza vaccines need to be updated every few years because of antigenic drift (2). Despite intensive monitoring, strain mismatch between vaccine formulation and influenza viruses circulating within the population has occurred in the past (2). Public health is further compromised when an unpredictable mixing event among influenza virus genomes leads to antigenic shift facilitating a potential pandemic outbreak. These concerns have expedited efforts toward developing a universal influenza vaccine.Neutralizing antibodies (nAbs) against hemagglutinin (HA) are the primary correlate for protection in humans and hence HA is an attractive target for vaccine development (3). The precursor polypeptide, HA0, is assembled into a trimer along the secretory pathway and transported to the cell surface. Cleavage of HA0 generates the disulfide-linked HA1 and HA2 subunits. Mature HA has a globular head domain which mediates receptor binding and is primarily composed of the HA1 subunit, whereas the stem domain predominantly comprises the HA2 subunit. The HA stem is trapped in a metastable state and undergoes an extensive low-pH-induced conformational rearrangement in the host-cell endosomes to adopt the virus–host membrane fusion-competent state (4, 5).The antigenic sites on the globular head of HA are subjected to heightened immune pressure resulting in escape variants, thereby limiting the breadth of head-directed nAbs (6). However, extensive efforts have resulted in the isolation of monoclonal antibodies (mAbs) that bind within the globular head and inhibit receptor attachment, which neutralize drifted variants of an HA subtype or heterosubtypic HA (7–16). The HA stem is targeted by several broadly neutralizing antibodies (bnAbs) with neutralizing activity against diverse influenza A virus subtypes (17). The epitopes of these bnAbs in the HA stem are more conserved across different influenza HA subtypes compared with the antigenic sites in the HA globular head (18).During a primary infection, the immunodominant globular head domain suppresses the response toward the conserved stem. Several efforts have been made to circumvent this problem. Repeated immunizations with full-length, chimeric HAs (cHAs) in a protracted vaccination regimen have been shown to boost stem-directed responses in mice (19). Alternatively, full-length HA presented on nanoparticles (np) has been shown to elicit stem-directed nAbs (20). Attempts have also been made to steer the immune response toward the conserved HA stem by hyperglycosylating the head domain (21). Although the aforementioned strategies need to be further evaluated and provide novel alternatives, detrimental interference from the highly variable immunodominant head domain in eliciting a broad functional response cannot be completely evaded. A “headless” stem domain immunogen offers an attractive solution. However, early attempts at expressing the HA2 subunit independently in a native, prefusion conformation were unsuccessful. In the absence of the head domain, the HA2 subunit expressed in Escherichia coli spontaneously adopted the low-pH conformation (22) in which the functional epitopes of stem-directed bnAbs are disrupted. More recently, the entire HA stem region has been expressed in a prefusion, native-like conformation in both prokaryotic and eukaryotic systems adopting multiple strategies (23–26).Design of independently folding HA stem fragments which adopt the prefusion HA conformation presents another approach to elicit bnAbs against influenza (27, 28). The A helix of the HA2 subunit contributes substantial contact surface to the epitope of stem-directed bnAbs such as CR6261, F10, and others. Although multivalent display of A helix on the flock house virus as a virus-like particle platform elicited cross-reactive antibodies, it conferred only minimal protection (20%) against virus challenge in mice (29).We report the design and characterization of engineered headless HA stem immunogens based on the influenza A/Puerto Rico/8/34 (H1N1) subtype. H1HA10-Foldon, a trimeric derivative of our parent construct (H1HA10), bound conformation-sensitive, stem-directed bnAbs such as CR6261 (30), F10 (31), and FI6v3 (32) with a high-affinity [equilibrium dissociation constant (K_D) of 10–50 nM]. The designed immunogens elicited broadly cross-reactive antiviral antibodies which neutralized highly drifted influenza virus strains belonging to both group 1 (H1 and H5 subtypes) and 2 (H3 subtype) in vitro. Significantly, stem immunogens designed from unmatched, highly drifted influenza strains conferred protection against a lethal (2LD₉₀) heterologous A/Puerto Rico/8/34 virus challenge in mice. Our immunogens confer robust subtype-specific and modest heterosubtypic protection in vivo. In contrast to previous stem domain immunogens (23–25), the designed immunogens were purified from the soluble fraction in E. coli. The HA stem-fragment immunogens do not aggregate even at high concentrations and are cysteine-free, which eliminates the complications arising from incorrect disulfide-linked, misfolded conformations. The aforementioned properties of the HA stem-fragment immunogens make it amenable for scalability at short notice which is vital during pandemic outbreaks.     

Drug Discovery Research in India: Current State and Future Prospects

Indian civilization developed a strong system of traditional medicine and was one of the first nations to develop a synthetic drug. In the postindependence era, Indian pharmaceutical industry developed a strong base for production of generic drugs. Challenges for the future are to give its traditional medicine a strong scientific base and develop research and clinical capability to consistently produce new drugs based on advances in modern biological sciences.Indian civilization is one of the few in the world that developed a full-fledged system of traditional medicine. The approach of Indian traditional medicine, e.g., the ayurvedic system, is herbal based in general and is more effective for chronic diseases and prevention. Although modern medicine has found its own niche in India, traditional formulations are still widely used, and more and more scientifically validated formulations are appearing in the market. In recent times, many plants used in Indian system of medicine have been analyzed by modern analytical methods and active components have been isolated. Significant amount of medicinal chemistry efforts are going on around these molecules in an attempt to develop more potent leads. These include curcumin from turmeric,¹ Bacosides from Brahmi (Bacopa monnieri),² and Forskolin from Coleus forskohlii. The first modern synthetic drug to be developed in India was Urea Stibamine in 1922 by UN Brahmachari against visceral leishmaniasis.³ Visceral leishmaniasis was a severe health burden during the early part of the 20th century, and it was a life saving drug for a large section of the population. Historically, it was the second drug developed against an infectious disease after Salversan (against Syphillis) and well before penicillin or sulfa drugs. It is still in use in many countries in a modified form.     

Bilateral luteomas of pregnancy in a patient with diabetes

A patient with diabetes and luteomas of the ovaries demonstrated a significant and progressive rise in the serum levels of androstenedione and testosterone during pregnancy. The levels of these hormones were also elevated in cord blood and ovarian fluid. The increased insulin requirement to maintain euglycemia was observed at an earlier date in this pregnancy than in her previous pregnancy.     

Abnormalities in metabolic pathways in celiac disease investigated by the metabolic profiling of small intestinal mucosa,blood plasma and urine by NMR spectroscopy

Celiac disease (CeD) is an autoimmune enteropathy caused by gluten intake in genetically predisposed individuals. We investigated the metabolism of CeD by metabolic profiling of intestinal mucosa, blood plasma and urine using NMR spectroscopy and multivariate analysis. The metabolic profile of the small intestinal mucosa was compared between patients with CeD (n = 64) and disease controls (DCs, n = 30). The blood plasma and urinary metabolomes of CeD patients were compared with healthy controls (HCs, n = 39). Twelve metabolites (proline (Pro), arginine (Arg), glycine (Gly), histidine (His), glutamate (Glu), aspartate, tryptophan (Trp), fumarate, formate, succinate (Succ), glycerophosphocholine (GPC) and allantoin (Alln)) of intestinal mucosa differentiated CeD from controls. The metabolome of blood plasma with 18 metabolites (Pro, Arg, Gly, alanine, Glu, glutamine, glucose (Glc), lactate (Lac), acetate (Ace), acetoacetate (AcAc), β‐hydroxybutyrate (β‐OHB), pyruvate (Pyr), Succ, citrate (Cit), choline (Cho), creatine (Cr), phosphocreatine (PCr) and creatinine) and 9 metabolites of urine (Pro, Trp, β‐OHB, Pyr, Succ, N‐methylnicotinamide (NMN), aminohippurate (AHA), indoxyl sulfate (IS) and Alln) distinguished CeD from HCs. Our data demonstrated changes in nine metabolic pathways. The altered metabolites were associated with increased oxidative stress (Alln), impaired healing and repair mechanisms (Pro, Arg), compromised anti‐inflammatory and cytoprotective processes (Gly, His, NMN), altered energy metabolism (Glc, Lac, β‐OHB, Ace, AcAc, Pyr, Succ, Cit, Cho, Cr and PCr), impaired membrane metabolism (GPC and Cho) and intestinal dysbiosis (AHA and IS). An orthogonal partial least square discriminant analysis model provided clear differentiation between patients with CeD and controls in all three specimens. A classification model built by combining the distinguishing metabolites of blood plasma and urine samples gave an AUC of 0.99 with 97.7% sensitivity, 93.3% specificity and a predictive accuracy of 95.1%, which was higher than for the models built separately using small intestinal mucosa, blood plasma and urine. In conclusion, a panel of metabolic biomarkers in intestinal biopsies, plasma and urine samples has potential to differentiate CeD from controls and may complement traditional tests to improve the diagnosis of CeD.     

Diffusion MRI detects early brain microstructure abnormalities in 2‐month‐old 3×Tg‐AD mice

The 3×Tg‐AD mouse is one of the most studied animal models of Alzheimer's disease (AD), and develops both amyloid beta deposits and neurofibrillary tangles in a temporal and spatial pattern that is similar to human AD pathology. Additionally, abnormal myelination patterns with changes in oligodendrocyte and myelin marker expression are reported to be an early pathological feature in this model. Only few diffusion MRI (dMRI) studies have investigated white matter abnormalities in 3×Tg‐AD mice, with inconsistent results. Thus, the goal of this study was to investigate the sensitivity of dMRI to capture brain microstructural alterations in 2‐month‐old 3×Tg‐AD mice. In the fimbria, the fractional anisotropy (FA), kurtosis fractional anisotropy (KFA), and radial kurtosis (K_┴) were found to be significantly lower in 3×Tg‐AD mice than in controls, while the mean diffusivity (MD) and radial diffusivity (D_┴) were found to be elevated. In the fornix, K_┴ was lower for 3×Tg‐AD mice; in the dorsal hippocampus MD and D_┴ were elevated, as were FA, MD, and D_┴ in the ventral hippocampus. These results indicate, for the first time, dMRI changes associated with myelin abnormalities in young 3×Tg‐AD mice, before they develop AD pathology. Morphological quantification of myelin basic protein immunoreactivity in the fimbria was significantly lower in the 3×Tg‐AD mice compared with the age‐matched controls. Our results demonstrate that dMRI is able to detect widespread, significant early brain morphological abnormalities in 2‐month‐old 3×Tg‐AD mice.