Addressing Reemergence of Diphtheria among Adolescents through Program Integration in India.

We report a diphtheria outbreak mostly among children (median 12 years; range 4-26 years) of a religious minority in urban India. Case-fatality rate (15%, 19/124) was higher among unimmunized patients (relative risk 4.1, 95% CI 1.5-11.7). We recommend mandating and integrating immunization into school health programs to prevent reemergence.

Differential levels of CHMP2B, LLPH, and SLC25A51 proteins in secondary renal amyloidosis.

OBJECTIVES: Renal amyloidosis (RA) is a rare disease, typically manifested with proteinuria, nephrotic syndrome, and ultimately leads to renal failure. The present study aims to profile the proteomes of renal amyloidosis patient's serum and healthy controls, along with relative quantification to find out robust markers for RA. METHODS: In this study, 12 RA patients and their corresponding age and gender-matched healthy controls were recruited from the Nephrology department of Max Super Specialty Hospital, New Delhi. We employed gel-based proteomic approach coupled with MALDI-TOF MS to compare protein expression patterns in RA patients and controls. Furthermore, validation of differential proteins (selected) was done using bio-layer interferometry. RESULTS: Eleven proteins showed remarkably altered expression levels. Moreover, expression modulation of three proteins (LLPH, SLC25A51, and CHMP2B) was validated which corroborated with two-dimensional gel electrophoresis (2-DE) results showing significant upregulation (p < 0.05) in RA patients followed by ROC analysis which demonstrated the diagnostic potential of these proteins. A protein-protein master network was generated implicating the above identified proteins along with their interactors, fishing out the routes leading to amyloidosis. CONCLUSION: This study indicates that the identified serum proteomic signatures could improve early diagnosis and lead to possible therapeutic targets in RA.

Particle encapsulation in aqueous ferrofluid drops and sorting of particle-encapsulating drops from empty drops using a magnetic field.

Handling and manipulation of particle-encapsulating droplets (PED) have profound applications in biochemical assays. Herein we report encapsulation of microparticles in aqueous ferrofluid droplets in a primary continuous phase (CP) and sorting of PED from empty droplets (ED) at the interface of the CP in coflow with a second continuous phase using a magnetic field. We find that the encapsulation process results in a size contrast between the PED and ED that depends on the flow regime - squeezing, dripping, or jetting - which in turn is governed by the ratio of the discrete phase to the continuous phase capillary number, Car. The difference between the volume fractions of ferrofluid in the PED and ED, ΔαPED, is utilized for sorting, and is found to depend on the ratio of the capillary numbers, Car. The difference ΔαPED is found to be maximum in the jetting regime, suggesting that the jetting regime is most suitable for encapsulation and sorting. The sorting criterion is represented in terms of a parameter ξ, which is a function of the ratios of the magnetic force to the interfacial force experienced by the PED and ED. Our study revealed that sorting is possible for ξ < 0, which corresponds to ΔαPED > 0.25. The maximum sorting efficiency of our system is found to be ∼95% at a throughput of ∼100 drops per s.

Differential expression of Lumican, Mimecan, Annexin A5 and Serotransferrin in ectopic and matched eutopic endometrium in ovarian endometriosis: a case-control study.

AIM: Endometriosis is a debilitating disease marked by recurrent gynecological proliferations. The present study aimed at performing differential proteomic analysis of matched eutopic and ectopic endometrium from women with ovarian endometriosis. MATERIALS AND METHODS: Proteomes were resolved using nano LC-MS and further identified and quantified using ProteinLynx Global SERVER (PLGS) software. Selected proteins were further chosen for validation by real time-polymerase chain reaction (RT-PCR). RESULTS: The protein profiles uncovered several differentially expressed proteins in the diseased sample (ectopic endometrium) as compared to the reference sample (eutopic endometrium). The study involved an advanced proteomic approach, nano LC-MS, and validates for the first time the upregulation of Mimecan and Lumican proteins in endometriosis. CONCLUSIONS: These proteins may hence prove as potentially useful tools in the search for diagnostic markers for early detection of the disease.

Fluoride Geochemistry and Exposure Risk Through Groundwater Sources in Northeastern Parts of Rajasthan, India.

Exposure to fluoride concentrations above a threshold of 1.5 mg/L can cause joint pains, restricted mobility, skeletal and dental fluorosis. This study aims to determine the hydrochemical evolution of the fluoride-rich groundwater and estimate the risk of fluoride exposure to the residents of semi-arid northeastern part of Rajasthan, India. The methodology involves measurement of fluoride and other ionic concentrations in groundwater using ion chromatography, followed by an estimation of the cumulative density function and fluorosis risk. The fluoride concentration in water samples varied from 0.04 to 8.2 mg/L with 85% samples falling above the permissible limit. The empirical cumulative density function was used to estimate the percentage and degree of health risks associated with the consumption of F- contaminated water. It is found that 55% of the samples indicate risk of dental fluorosis, 42% indicate risk of deformities to knee and hip bones, and 18% indicate risk of crippling fluorosis. In addition, instances of high nitrate concentrations above the permissible limit of 45 mg/L are also found in 13% of samples. The fluoride rich groundwater is mainly associated with the Na-HCO3-Cl type water facies while low fluoride groundwater shows varied chemical facies. The saturation index values indicate a high probability of a further increase in F- concentration in groundwater of this region. The calculated fluoride exposure risk for the general public in the study area is 3-6 times higher than the allowed limit of 0.05 mg/kg/day. Based on the results of this study, a fluorosis index map was prepared for the study area. The northern and northeastern parts are less prone to fluorosis, whereas the south-central and southwestern parts are highly vulnerable to fluorosis. The inferences from this study help to prioritize the regions that need immediate attention for remediation.

Differential expression of MAP3K7 and TROPONIN C proteins and related perturbations in renal amyloidosis.

OBJECTIVES: Renal amyloidosis (RA) is a rare protein misfolding disorder that prompts progressive renal insufficiency. This study aimed to decipher proteomic changes in human sera to understand the pathophysiology and molecular mechanisms underlying the disease development, hence assisting in the diagnosis of RA. METHODS: Serum proteomic analysis was performed using a gel-based approach followed by MALDI-TOF MS. RA patients with age and sex matched healthy volunteers were recruited from Max Super Speciality Hospital, New Delhi, India. RESULTS: Proteome profiles of serum revealed eight differentially expressed proteins namely, Zinc finger protein 624, Protein FAM183A, Calcium-binding mitochondrial carrier protein Scamc-3, V-type proton ATPase 116 kDa subunit A isoforms 2, Protein TXNRD3NB, ATP - dependent RNA helicase, Troponin C and Mitogen-activated protein kinase kinase kinase 7. These proteins were reported first time in RA. The increased levels of MAP3K7 and TROPONIN C were validated by bio-layer interferometry and their diagnostic accuracy was evaluated by ROC curve analysis. The differentially expressed proteins were predominantly associated with vesicular trafficking, transcriptional regulation, metabolic processes, apoptotic process and mitochondrial metabolism. CONCLUSION: The results indicate that these proteomic signatures may be considered as potential molecular targets for RA diagnostics and therapeutics subject to validation on large sample size. Abbreviations: AßP= Amyloid-beta protein, Aß=Amyloid-beta, AL= Light chain amyloidosis, AA= Amyloid A, ALECT2= LECT2 amyloidosis, APS= Ammonium persulfate CKD= Chronic Kidney Diseases, EBRT= external beam radiation therapy, ESRD= End-Stage Kidney Disease, Glis2= Gli-similar 2, JNK= c-Jun NH 2-terminal kinase, MAPK= Mitogen-Activated Protein Kinase, MM=Multiple Myeloma, PHD= Prolyl hydroxylase, RA = Renal Amyloidosis, SAA= Serum Amyloid A, SD= Standard Deviation, Sepp= Selenoprotein, SCC= Squamous cell carcinoma, SDS= Sodium dodecyl sulfate, TEMED = tetramethyl ethylenediamine, TGF-Beta-1=Transforming growth factor- Beta-1, Trx = Thioredoxin, TrxR= Thioredoxin reductase.

Proteomic sift through serum and endometrium profiles unraveled signature proteins associated with subdued fertility and dampened endometrial receptivity in women with polycystic ovary syndrome.

The objective of this study is to discern the proteomic differences responsible for hampering the receptivity of endometrium and subduing the fertility of females with polycystic ovary syndrome in analogy to healthy fertile females. This study was designed in collaboration with Hakeem Abdul Hameed Centenary Hospital affiliated to Jamia Hamdard, New Delhi, India. Serum samples were taken from infertile PCOS subjects (n = 6) and fertile control subjects (n = 6) whereas endometrial tissue samples were recruited from ovulatory PCOS (n = 4), anovulatory PCOS (n = 4) and normal healthy fertile control subjects (n = 4) for proteomic studies. Additionally, endometrial biopsies from healthy fertile control (n = 8), PCOS with infertility (n = 6), unexplained infertility (n = 3) and endometrial hyperplasia (n = 3) were taken for validation studies. Anthropometric, biochemical and hormonal evaluation was done for all the subjects enrolled in this study. Protein profiles were generated through 2D-PAGE and differential proteins analyzed with PD-QUEST software followed by identification with MALDI-TOF MS protein mass fingerprinting. Validation of identified proteins was done through RT-PCR relative expression analysis. Protein profiling of serum revealed differential expression of proteins involved in transcriptional regulation, embryogenesis, DNA repair, decidual cell ploidy, immunomodulation, intracellular trafficking and degradation processes. Proteins involved in cell cycle regulation, cellular transport and signaling, DNA repair, apoptotic processes and mitochondrial metabolism were found to be differentially expressed in endometrium. The findings of this study revealed proteins that hold strong candidature as potential drug targets to regulate the cellular processes implicating infertility and reduced receptivity of endometrium in women with polycystic ovary syndrome.

Trapping and Coalescence of Diamagnetic Aqueous Droplets Using Negative Magnetophoresis.

The manipulation of aqueous droplets has a profound significance in biochemical assays. Magnetic field-driven droplet manipulation, offering unique advantages, is consequently gaining attention. However, the phenomenon relating to diamagnetic droplets is not well understood. Here, we report the understanding of trapping and coalescence of flowing diamagnetic aqueous droplets in a paramagnetic (oil-based ferrofluid) medium using negative magnetophoresis. Our study revealed that the trapping phenomenon is underpinned by the interplay of magnetic energy (Em) and frictional (viscous) energy (Ef), in terms of magnetophoretic stability number, Sm = (Em/Ef). The trapping and nontrapping regimes are characterized based on the peak value of magnetophoretic stability number, Smp, and droplet size, D*. The study of coalescence of a trapped droplet with a follower droplet (and a train of droplets) revealed that the film-drainage Reynolds number (Refd) representing the coalescence time depends on the magnetic Bond number, Bom. The coalesced droplet continues to remain trapped or gets self-released obeying the Smp and D* criterion. Our study offers an understanding of the magnetic manipulation of diamagnetic aqueous droplets that can potentially be used for biochemical assays in microfluidics.

Assessment of acetamiprid-induced genotoxic effects in bone marrow cells of Swiss albino male mice.

Acetamiprid (ACE), a neonicotinoid insecticide, is widely used in agriculture either alone or in combination with other insecticides. A combined approach employing micronucleus test (MNT) and chromosomal aberrations (CA) assay was utilized to assess the genotoxic effects of ACE in bone marrow of Swiss albino male mice. Acetamiprid was administered i.p. daily at 4.6 and 2.3 mg/kg/day along with 3% gum acacia as negative control for 60 and 90 days and cyclophosphamide (50 mg/kg b.wt.) as positive control. ACE treatment resulted in a dose-dependent increase in the frequencies of micronuclei per cell and chromosomal aberrations in bone marrow cells. The increased micronuclei formation in total erythrocyte cells (immature PCEs and mature NCEs) was observed only at higher dose level (4.6 mg/kg b.wt.) administered for 90 days. The test also indicated the cytotoxic effect of higher dose level of pesticide by PCE/NCE ratio. The number of chromosomal aberrations were increased in the pesticide treated group compared to the negative control group, although significant increase was observed only in the group exposed to higher dose level of pesticide for both 60 and 90 days. Thus, daily exposure of ACE at a dose level of 4.6 mg/kg body weight for 60 and 90 days caused genotoxic and cytotoxic effects on the somatic cells of Swiss albino male mice.

Association of IL-1ß, IL-1Ra and FABP1 gene polymorphisms with the metabolic features of polycystic ovary syndrome.

BACKGROUND: Polycystic ovary syndrome (PCOS), a highly prevalent endocrinopathy is currently being designated as chronic low grade inflammatory state. IL-1ß, IL-1Ra and FABP1 are critical mediators of inflammatory processes and are speculated to play a role in the pathogenesis of PCOS. The aim of this study was to study the association of IL-ß, IL-1Ra and FABP1 gene polymorphisms with PCOS and related metabolic features. SUBJECTS: 95 PCOS and 45 age matched healthy control subjects were enrolled in this study. METHODS: Polymorphism in genes IL-1ß, IL-1Ra and FABP1 was studied by PCR, PCR-RFLP and sequencing methods, respectively. Hormonal and lipid profiles were evaluated for all the subjects. RESULTS: Hormonal and lipid profiles showed significant differences between PCOS and control subjects. Allele and genotype frequencies of IL-1ß, IL-1Ra and FABP1 gene polymorphisms did not vary between the control and PCOS group. However, T allele of C[-511]T variant of IL-1ß, allele II in intron 2 of IL-1Ra and A allele of A/G variant of FABP1 (rs2197076) showed significant association with many metabolic features associated with PCOS. CONCLUSIONS: Polymorphism in genes encoding cytokines and proteins involved in lipid metabolism can provide insights into the genetics of the disease and may contribute to assess the associated risk of cardiovascular diseases (CVD), dyslipidemia and metabolic syndrome (MetS) associated with PCOS.

Non-native ligands define the active site of Pennisetum glaucum (L.) R. Br dehydroascorbate reductase.

Dehydroascorbate reductase (DHAR), a member of the glutathione-S-transferase (GST) family, reduces dehydroascorbate (DHA) to ascorbate (AsA; Vitamin-C) in a glutathione (GSH)-dependent manner and in doing so, replenishes the critical AsA pool of the cell. To understand the enzyme mechanism in detail, we determined the crystal structure of a plant DHAR from Pennisetum glaucum (PgDHAR) using Iodide-Single Anomalous Dispersion (SAD) and Molecular replacement methods, in two different space groups. Here, we show PgDHAR in complex with two non-native ligands, viz. an acetate bound at the G-site, which resembles the γ-carboxyl moiety of GSH, and a glycerol at the H-site, which shares the backbone of AsA. We also show that, in the absence of bound native substrates, these non-native ligands help define the critical 'hook points' in the DHAR enzyme active site. Further, our data suggest that these non-native ligands can act as the logical bootstrapping points for iterative design of inhibitors/analogs for DHARs.

Serum microRNA-21 expression as a prognostic and therapeutic biomarker for breast cancer patients.

MiRNA-21 is recognized as the main active candidate and high expression in many solid tumors consequential cell proliferation, differentiation, apoptosis, and closely related to metastasis of disease. The study aimed to evaluate the serum miRNA-21 expression and therapy outcome in breast cancer patients and cell lines. Seventy-five histopathologically confirmed newly diagnosed breast cancer patients were included in the study; before and after therapy, patient's blood sample were collected and analyzed for serum microRNA-21 expression by quantitative real-time PCR. In patients, 8.9 mean fold increased microRNA-21 expression was observed compared to controls. Increased expression was found to be associated with advanced stage (11.72-fold), lymph node involvement (11.12-fold), and distant metastases (20.17-fold). After treatment significant decrease in miRNA-21 expression was observed and found to be significant (p < 0.0001). Patients treated with neoadjuvant therapy had significant impact on miRNA-21 suppression and found to be significantly associated with different clinicopathological features of patients. Increased miRNA-21 expression was also found to be significantly associated with poor survival of breast cancer patients (p = 0.002). MicroRNA-21 expression could be used as promising predictive indicators for breast cancer prognosis. MicroRNA-21 over-expression was associated with response to neoadjuvant therapy and may perhaps be considered as primary treatment choice.

Development of a recombinant hCG-specific single chain immunotoxin cytotoxic to hCG expressing cancer cells.

A large number of cancers express human chorionic gonadotropin (hCG) or its subunits ectopically. Patients harboring such cancers have poor prognosis and adverse survival. PiPP is a monoclonal antibody of high affinity and specificity for hCGß/hCG. Work was carried out to develop a PiPP based recombinant immunotoxin for the immunotherapy of hCG expressing cancers. Recombinant PiPP antibody was constructed in scFv format in which gene encoding the VH and VL domains were joined through a linker. This scFv gene was fused to the gene expressing Pseudomonas exotoxin (PE38), and cloned in a Escherichia coli based expression vector under the control of strong bacteriophage T7 promoter. Immunotoxin conjugating scFv(PiPP) and PE38, was expressed in E. coli as recombinant protein. Recombinant PiPP immunotoxin was purified from the bacterial cell lysate and tested for binding and killing of hCGß expressing lymphoma, T-lymphoblastic leukemia and lung carcinoma cells in vitro. Immunotoxin showed nearly 90% killing on the cells. This is the first ever report on recombinant immunotoxin for binding and cytotoxicity to hCG expressing cancer cells, and thus can be a potential candidate for the immunotherapy of hCG expressing cells.

Unusual co-existence of dural sinus thrombosis and aneurysmal subarachnoid hemorrhage in a patient with lupus.

Subarachnoid hemorrhage and dural sinus thrombosis are important manifestations of neuropsychiatric lupus erythematosus. We report the case of a woman with relapsed lupus nephritis, in partial remission, who presented with the unusual combination of dural sinus thrombosis (due to protein S functional deficiency) and aneurysmal subarachnoid hemorrhage. She had a fatal outcome, as has often been reported with subarachnoid hemorrhage in the context of active lupus. To our knowledge, this is the first report of such an unusual association in lupus, and highlights unique challenges in the management of intracranial hemorrhage in the context of dural sinus thrombosis.

Eradicating successfully yaws from India: The strategy & global lessons.

Yaws, a non-venereal treponematosis, affecting primarily the tribal populations, has been considered historically as one of the most neglected tropical diseases in the world. In 1996, India piloted an initiative to eradicate yaws based on a strategy consisting of active case finding through house-to-house search and treatment of cases and their contacts with long acting penicillin. Thereafter, the campaign implemented in all 51 endemic districts in 10 states of the country led to the achievement of a yaws-free status in 2004. In the post-elimination phase, surveillance activities accompanied by serological surveys were continued in the erstwhile endemic districts. These surveys carried out among children between the age of 1-5 yr, further confirmed the absence of community transmission in the country. The experience of India demonstrates that yaws can be eradicated in all endemic countries of Africa and Asia, provided that political commitment can be mobilized and community level activities sustained until the goal is achieved.

Structural characteristics of thermostable immunogenic outer membrane protein from Salmonella enterica serovar Typhi.

In this work, we explored the acid-induced unfolding pathway of non-porin outer membrane protein (OMP), an immunogenic protein from Salmonella Typhi, by monitoring the conformational changes over a pH range of 1.0-7.0 by circular dichroism, intrinsic fluorescence, ANS binding, acrylamide quenching, and dynamic light scattering. The spectroscopic measurements showed that OMP in its native state at pH 7.0 exists in more stable and compact conformation. In contrast, at pH 2.0, OMP retains substantial amount of secondary structure, disrupted side chain interactions, increased hydrodynamic radii, and nearly four-fold increase in ANS fluorescence with respect to the native state, indicating that MG state exists at pH 2.0. Quenching of tryptophan fluorescence by acrylamide further confirmed the accumulation of a partially unfolded state between native and unfolded state. The effect of pH on the conformation and thermostability of OMP points towards its heat resistance at neutral pH (T m ~ 69 °C at pH 7.0, monitored by change in MRE222 nm). Acid unfolded state was also characterized by the lack of a cooperative thermal transition. All these results suggested that acid-induced unfolded state of OMP at pH 2.0 represented the molten globule state. The chemical denaturation studies with GuHCl and urea as denaturants showed dissimilar results. The chemical unfolding experiments showed that in both far-UV CD and fluorescence measurements, GuHCl is more efficient than urea. GuHCl is characterized by low C m (~1 M), while urea is characterized by high C m (~3 M). The fully unfolded states were reached at 2 M GuHCl and 4 M urea concentration, respectively. This study adds to several key considerations of importance in the development of therapeutic agents against typhoid fever for clinical purposes.

Leucoagglutinating phytohemagglutinin: purification, characterization, proteolytic digestion and assessment for allergenicity potential in BALB/c mice.

Red kidney bean (Phaseolus vulgaris) is consumed worldwide as a vegetarian protein source. But, at the same time the allergenicity potential of red kidney bean is a matter of concern. This study is aimed towards purification, characterization, thermal stability, proteolytic digestion and allergenicity assessment of one of the clinically relevant allergens of red kidney bean. The purification of red kidney bean allergic protein was carried out with the help of column chromatography, IgE immunoblotting and reverse phase high-pressure liquid chromatography (RP-HPLC). The purified protein was characterized by peptide mass finger printing (PMF) and studied for its thermal stability, and proteolytic resistance using simulated gastric fluid (SGF) assay. The allergenicity potential of the purified protein was studied in BALB/c mice. The purified protein was identified as leucoagglutinating phytohemagglutinin (PHA-L) with molecular weight 29.5 kDa. The PHA-L showed resistance to heat as well as proteolytic enzyme. Higher levels of total IgE, specific IgE, and histamine were observed in PHA-L treated BALB/c mice when compared to control. Overall, PHA-L possesses characteristics of allergens and may play a potential role in the red kidney bean induced allergy.

The effect of a short-term pulmonary rehabilitation on exercise capacity and quality of life in patients hospitalised with acute exacerbation of chronic obstructive pulmonary disease.

BACKGROUND: Recent research shows that pulmonary rehabilitation (PR) programmes in patients with acute exacerbation of chronic obstructive pulmonary disease (AECOPD), reduced dyspnoea, improved exercise capacity, and prevented occurrence of further exacerbations. OBJECTIVE: To evaluate the utility of a 3-week PR programme in patients with AECOPD. METHODS: Patients admitted with AECOPD, following clinical stabilisation in the respiratroy intensive care unit (RICU), were alternately assigned to intervention (n=15); and control groups (n=15), respectively. Baseline assessment included spirometry, six-minute walk test (6MWT), symptom limited cardiopulmonary exercise test (CPET), health-related quality of life (HRQoL) assessment by generic questionnaire medical outcomes study short form (S-F 36) questionnaire and dyspnoea evaluation by Borg score. The intervention group patients were treated with usual care plus PR exercises in the form of 20 minutes each of walking, bicycle ergometry and resistance exercises, thrice-weekly for three weeks. The control group patients were treated with only the usual care. After discharge from hospital the treatment regimens were continued on alternate days on outpatient basis, for a total of three weeks. The assessment was repeated in both the groups after three weeks. RESULTS: Nine sessions of PR exercises produced statistically significant improvement in general well-being, forced expiratory volume in the first second (FEV1), 6MWT parameters, exercise capacity, peak oxygen uptake and volume of oxygen consumption (VO2)/Watts slope on CPET in patients with AECOPD. CONCLUSION: Short duration PR programmes appear to be helpful in the management of AECOPD.