Implementing the WHO Labour Care Guide to reduce the use of Caesarean section in four hospitals in India: protocol and statistical analysis plan for a pragmatic, stepped-wedge, cluster-randomized pilot trial.

BACKGROUND: The World Health Organization (WHO) Labour Care Guide (LCG) is a paper-based labour monitoring tool designed to facilitate the implementation of WHO's latest guidelines for effective, respectful care during labour and childbirth. Implementing the LCG into routine intrapartum care requires a strategy that improves healthcare provider practices during labour and childbirth. Such a strategy might optimize the use of Caesarean section (CS), along with potential benefits on the use of other obstetric interventions, maternal and perinatal health outcomes, and women's experience of care. However, the effects of a strategy to implement the LCG have not been evaluated in a randomised trial. This study aims to: (1) develop and optimise a strategy for implementing the LCG (formative phase); and (2) To evaluate the implementation of the LCG strategy compared with usual care (trial phase). METHODS: In the formative phase, we will co-design the LCG strategy with key stakeholders informed by facility assessments and provider surveys, which will be field tested in one hospital. The LCG strategy includes a LCG training program, ongoing supportive supervision from senior clinical staff, and audit and feedback using the Robson Classification. We will then conduct a stepped-wedge, cluster-randomized pilot trial in four public hospitals in India, to evaluate the effect of the LCG strategy intervention compared to usual care (simplified WHO partograph). The primary outcome is the CS rate in nulliparous women with singleton, term, cephalic pregnancies in spontaneous labour (Robson Group 1). Secondary outcomes include clinical and process of care outcomes, as well as women's experience of care outcomes. We will also conduct a process evaluation during the trial, using standardized facility assessments, in-depth interviews and surveys with providers, audits of completed LCGs, labour ward observations and document reviews. An economic evaluation will consider implementation costs and cost-effectiveness. DISCUSSION: Findings of this trial will guide clinicians, administrators and policymakers on how to effectively implement the LCG, and what (if any) effects the LCG strategy has on process of care, health and experience outcomes. The trial findings will inform the rollout of LCG internationally. TRIAL REGISTRATION: CTRI/2021/01/030695 (Protocol version 1.4, 25 April 2022).

The new WHO Labour Care Guide (LCG) is an innovative partograph that emphasises women-centred, evidence-based care during labour and childbirth. Together with clinicians working at four hospitals in India, we will develop and test a strategy to implement the LCG into routine care in labour wards of these hospitals. We will use a randomised trial design where this LCG strategy is introduced sequentially in each of the four hospitals, in a random order. We will collect data on all women giving birth and their newborns during this period and analyse whether the LCG strategy has any effects on the use of Caesarean section, women's and newborn's health outcomes, and women's experiences during labour and childbirth. While the trial is being conducted, we will also collect qualitative and quantitative data from doctors, nurses and midwives working in these hospitals, to understand their perspectives and experiences of using the LCG in their day-to-day work. In addition, we will collect economic data to understand how much the LCG strategy costs, and how much money it might save if it is effective. Through this study, our international collaboration will generate critical evidence and innovative tools to support implementation of the LCG in other countries.

Two Phase Transitions in the Two-Dimensional Nematic Three-Vector Model with No Quasi-Long-Range Order: Monte Carlo Simulation of the Density of States.

The presence of stable topological defects in a two-dimensional (d=2) liquid crystal model allowing molecular reorientations in three dimensions (n=3) was largely believed to induce a defect-mediated Berzenskii-Kosterlitz-Thouless-type transition to a low temperature phase with quasi-long-range order. However, earlier Monte Carlo (MC) simulations could not establish certain essential signatures of the transition, suggesting further investigations. We study this model by computing its equilibrium properties through MC simulations, based on the determination of the density of states of the system. Our results show that, on cooling, the high temperature disordered phase deviates from its initial progression towards the topological transition, crossing over to a new fixed point, condensing into a nematic phase with exponential correlations of its director fluctuations. The thermally induced topological kinetic processes continue, however, limited to the length scales set by the nematic director fluctuations, and lead to a second topological transition at a lower temperature. It is argued that in the (d=2, n=3) system with an attractive biquadratic Hamiltonian, the presence of additional molecular degrees of freedom and local Z_{2} symmetry associated with lattice sites together promote the onset of an additional relevant scaling field at matching length scales in the high temperature region, leading to a crossover.

Immune recognition of tumor-associated mucin MUC1 is achieved by a fully synthetic aberrantly glycosylated MUC1 tripartite vaccine.

The mucin MUC1 is typically aberrantly glycosylated by epithelial cancer cells manifested by truncated O-linked saccharides. The resultant glycopeptide epitopes can bind cell surface major histocompatibility complex (MHC) molecules and are susceptible to recognition by cytotoxic T lymphocytes (CTLs), whereas aberrantly glycosylated MUC1 protein on the tumor cell surface can be bound by antibodies to mediate antibody-dependent cell-mediated cytotoxicity (ADCC). Efforts to elicit CTLs and IgG antibodies against cancer-expressed MUC1 have not been successful when nonglycosylated MUC1 sequences were used for vaccination, probably due to conformational dissimilarities. Immunizations with densely glycosylated MUC1 peptides have also been ineffective due to impaired susceptibility to antigen processing. Given the challenges to immuno-target tumor-associated MUC1, we have identified the minimum requirements to consistently induce CTLs and ADCC-mediating antibodies specific for the tumor form of MUC1 resulting in a therapeutic response in a mouse model of mammary cancer. The vaccine is composed of the immunoadjuvant Pam(3)CysSK(4), a peptide T(helper) epitope and an aberrantly glycosylated MUC1 peptide. Covalent linkage of the three components was essential for maximum efficacy. The vaccine produced CTLs, which recognized both glycosylated and nonglycosylated peptides, whereas a similar nonglycosylated vaccine gave CTLs which recognized only nonglycosylated peptide. Antibodies elicited by the glycosylated tripartite vaccine were significantly more lytic compared with the unglycosylated control. As a result, immunization with the glycosylated tripartite vaccine was superior in tumor prevention. Besides its own aptness as a clinical target, these studies of MUC1 are likely predictive of a covalent linking strategy applicable to many additional tumor-associated antigens.

Myeloid-derived suppressor cells adhere to physiologic STAT3- vs STAT5-dependent hematopoietic programming, establishing diverse tumor-mediated mechanisms of immunologic escape.

The receptor tyrosine kinase inhibitor, sunitinib, is astonishingly effective in its capacity to reduce MDSCs in peripheral tissues such as blood (human) and spleen (mouse), restoring responsiveness of bystander T lymphocytes to TcR stimulation. Sunitinib blocks proliferation of undifferentiated MDSCs and decreases survival of more differentiated neutrophilic MDSC (n-MDSC) progeny. Ironically, sunitinib's profound effects are observed even in a total absence of detectable anti-tumor therapeutic response. This is best explained by the presence of disparate MDSC-conditioning stimuli within individual body compartments, allowing sensitivity and resistance to sunitinib to coexist within the same mouse or patient. The presence or absence of GM-CSF is likely the major determinant in each compartment, given that GM-CSF's capacity to preempt STAT3-dependent with dominant STAT5-dependent hematopoietic programming confers sunitinib resistance and redirects differentiation from the n-MDSC lineage to the more versatile monocytoid (m-MDSC) lineage. The clinical sunitinib experience underscores that strategies for MDSC and Treg depletions must be mindful of disparities among body compartments to avoid sanctuary effects. Ironically, m-MDSCs manifesting resistance to sunitinib also have the greatest potential to differentiate into tumoricidal accessory cells, by virtue of their capacity to respond to T cell-secreted IFN-γ or to TLR agonists with nitric oxide and peroxynitrate production.

Thirteen-week oral toxicity study of carotenoid pigment from Rhodotorula glutinis DFR-PDY in rats.

Carotenoids from some of the coloured yeasts like Rhodotorula, Phaffia rhodozyma have attracted commercial interest as a natural pigment for foods. Red yeast isolated from contaminated Potato dextrose agar plate (PDA), designated as Rhodotorula glutinis DFR-PDY has been found to produce carotenoids. In the present study toxicological evaluation of carotenoid pigment has been reported. Experiment was conducted on 3 groups of albino rats. One group with vehicle control (palm oil) and 2 groups with two different doses of red yeast pigment (lower and higher dose) were fed to rats (both male and female) by gavages for 13 weeks. Gain in body weight of rats and food consumption were monitored at regular intervals. Hematological studies revealed that there is no much difference in erythrocytes, packed cell volume, Mean corpuscular volume (MCV), Mean corpuscular haemoglobin concentration (MCHC), platelets and differential counts. Total leucocyte count (TLC) is less in case of higher dose group than the lower and control groups. Whereas, hemoglobin is more in case of higher dose than the lower dose group and least in control group. Even clinico-chemical parameters and urine analysis of vehicle control group and pigment fed rats revealed that there were no major differences between them as well as between two different genders of rats and also interaction between different doses and the genders. Histopathology of these experimental animals revealed that there are no major histological changes found between the groups. It may be concluded that the whole pigment extract from R. glutinis DFR-PDY may be used safely in food preparations as food colourant with an added benefit of antioxidant activity.

Study of Prevalence of Abnormal Pap Smear and its Associated Risk Factors in HIV Positive Women: A Cross-sectional Study.

Objective: 1. To determine the prevalence of pre malignant cervical lesions in HIV positive women using conventional Pap smear. 2. To determine the association between various risk factors in HIV positive women and abnormal cytology on Pap smear. Design: A cross-sectional study was conducted in Bangalore Medical College in which eligible HIV-positive women underwent Pap smear, human papillomavirus (HPV) testing and cervical biopsy. Methods: Retropositive women attending gynaec OPD during the study period were taken into the study after taking informed consent. Women who fulfilled the inclusion criteria were subjected to Pap smear. Bethesda system of classification was used for reporting the Pap smear. Women with abnormal Pap smear were further evaluated by HPV DNA testing and cervical biopsy. Results: Cervical cytology was abnormal in 30% of the HIV-positive women, out of which 10% had HSIL, 15% had LSIL and 5% had ASCUS. Age at first sexual intercourse < 17 years (p = 0.009), past H/O STI (p = 0.0001), women with husband's having multiple sexual partners (p = 0.0001), women with CD4 count < 350 cells/micro-litre (p = 0.0001) were significant risk factors associated with abnormal Pap smear. Conclusion: Invasive cervical cancer is considered a preventable disease because of its long preinvasive state. Therefore, screening for premalignant cervical lesions represents an opportunity to prevent women developing cervical carcinoma.

Collagen induced arthritis increases secondary metastasis in MMTV-PyV MT mouse model of mammary cancer.

BACKGROUND: Several studies have demonstrated that sites of chronic inflammation are often associated with the establishment and growth of various malignancies. A common inflammatory condition in humans is autoimmune arthritis (AA). Although AA and cancer are different diseases, many of the underlying processes that contribute to the disorders of the joints and connective tissue that characterize AA also affect cancer progression and metastasis. Systemically, AA can lead to cellular infiltration and inflammation of the lungs. Several studies have reported statistically significant risk ratios between AA and breast cancer. Despite this knowledge being available, there has been minimal research linking breast cancer, arthritis, and metastasis associated with breast cancer. Notably both diseases are extremely prevalent in older post-menopausal women. METHODS: To establish the novel link between arthritis induced inflammation and secondary metastasis associated with breast cancer, PyV MT mice that spontaneously develop mammary gland carcinoma were injected with Type II collagen (CII) to induce arthritis at 9 and 18 weeks of age for pre-metastatic and metastatic condition. The sites of secondary metastasis and the associated inflammatory microenvironment were evaluated. RESULTS: A significant increase in breast cancer-associated secondary metastasis to the lungs and bones was observed in the arthritic versus the non-arthritic PyV MT mice along with an increase in primary tumor burden. We report significant increases in the levels of interstitial cellular infiltrates and pro-inflammatory cytokines such as interleukin-17 (IL-17), interleukin-6 (IL-6), Pro- Matrix metallopeptidase 9 (Pro-MMP9), insulin like growth factor-II (GF-II) and macrophage colony stimulating factor (M-CSF) in the arthritic lung and bone milieu as well as in the circulation. These pro-inflammatory cytokines along with the inflammatory microenvironment may be the underlying factors facilitating tumor progression and metastasis in arthritic PyV MT mice. This was further substantiated by treatment with celecoxib, an anti-inflammatory drug + αIL-17 antibody that significantly reduced the secondary metastasis to lung and bone. CONCLUSIONS: The data generated not only reveal the underlying mechanism of high susceptibility to bone and lung metastasis in an arthritic condition but our combination therapies may lead to treatment modalities that will be capable of reducing tumor burden, and preventing relapse and metastasis in arthritic patients with breast cancer.

Topological phase transitions in two-dimensional bent-core liquid crystal models.

Two-dimensional liquid crystal (LC) models of interacting V-shaped bent-core molecules with two rigid rodlike identical segments connected at a fixed angle (θ=112^{∘}) are investigated. The model assigns equal biquadratic tensor coupling among constituents of the interacting neighboring molecules on a square lattice, allowing for reorientations in three dimensions (d=2, n=3). We find evidence of two temperature-driven topological transitions mediated by point disclinations associated with the three ordering directors, condensing the LC medium successively into uniaxial and biaxial phases. With Monte Carlo simulations, temperature dependencies of the system energy, specific heat, orientational order parameters, topological order parameters, and densities of unbound topological defects of the different ordering directors are computed. The high-temperature transition results in topological ordering of disclinations of the primary director, imparting uniaxial symmetry to the phase. The low-temperature transition precipitates simultaneous topological ordering of defects of the remaining directors, resulting in biaxial symmetry. The correlation functions, quantifying spatial variations of the orientational correlations of the molecular axes show exponential decays in the high-temperature (disordered) phase, and power-law decays in the low-temperature (biaxial) phase. Differing temperature dependencies of the topological parameters point to a significant degree of cross coupling among the uniaxial and biaxial tensors of interacting molecules. This simplified Hamiltonian leaves θ as the only free model parameter, and the system traces a θ-dependent trajectory in a plane of the phenomenological parameter space.

Multipeptide stimulated PBMCs generate TEM/TCM for adoptive cell therapy in multiple myeloma.

Multiple Myeloma (MM) patients suffer disease relapse due to the development of therapeutic resistance. Increasing evidence suggests that immunotherapeutic strategies can provide durable responses. Here we evaluate the possibility of adoptive cell transfer (ACT) by generating ex vivo T cells from peripheral blood mononuclear cells (PBMCs) isolated from MM patients by employing our previously devised protocols. We designed peptides from antigens (Ags) including cancer testis antigens (CTAs) that are over expressed in MM. We exposed PBMCs from different healthy donors (HDs) to single peptides. We observed reproducible Ag-specific cluster of differentiation 4+ (CD4+) and CD8+ T cell responses on exposure of PBMCs to different single peptide sequences. These peptide sequences were used to compile four different peptide cocktails. Naïve T cells from PBMCs from MM patients or HDs recognized the cognate Ag in all four peptide cocktails, leading to generation of multiclonal Ag-specific CD4+ and CD8+ effector and central memory T (TEM and TCM, respectively) cells which produced interferon-gamma (IFN-γ), granzyme B and perforin on secondary restimulation. Furthermore, this study demonstrated that immune cells from MM patients are capable of switching metabolic programs to induce effector and memory responses. Multiple peptides and cocktails were identified that induce IFN-γ+, T1-type, metabolically active T cells, thereby paving the way for feasibility testing of ACT in phase I clinical trials.

Factors associated with mortality among moderate and severe patients with COVID-19 in India: a secondary analysis of a randomised controlled trial.

OBJECTIVE: Large data on the clinical characteristics and outcome of COVID-19 in the Indian population are scarce. We analysed the factors associated with mortality in a cohort of moderately and severely ill patients with COVID-19 enrolled in a randomised trial on convalescent plasma. DESIGN: Secondary analysis of data from a Phase II, Open Label, Randomized Controlled Trial to Assess the Safety and Efficacy of Convalescent Plasma to Limit COVID-19 Associated Complications in Moderate Disease. SETTING: 39 public and private hospitals across India during the study period from 22 April to 14 July 2020. PARTICIPANTS: Of the 464 patients recruited, two were lost to follow-up, nine withdrew consent and two patients did not receive the intervention after randomisation. The cohort of 451 participants with known outcome at 28 days was analysed. PRIMARY OUTCOME MEASURE: Factors associated with all-cause mortality at 28 days after enrolment. RESULTS: The mean (SD) age was 51±12.4 years; 76.7% were males. Admission Sequential Organ Failure Assessment score was 2.4±1.1. Non-invasive ventilation, invasive ventilation and vasopressor therapy were required in 98.9%, 8.4% and 4.0%, respectively. The 28-day mortality was 14.4%. Median time from symptom onset to hospital admission was similar in survivors (4 days; IQR 3-7) and non-survivors (4 days; IQR 3-6). Patients with two or more comorbidities had 2.25 (95% CI 1.18 to 4.29, p=0.014) times risk of death. When compared with survivors, admission interleukin-6 levels were higher (p<0.001) in non-survivors and increased further on day 3. On multivariable Fine and Gray model, severity of illness (subdistribution HR 1.22, 95% CI 1.11 to 1.35, p<0.001), PaO2/FiO2 ratio <100 (3.47, 1.64-7.37, p=0.001), neutrophil lymphocyte ratio >10 (9.97, 3.65-27.13, p<0.001), D-dimer >1.0 mg/L (2.50, 1.14-5.48, p=0.022), ferritin ≥500 ng/mL (2.67, 1.44-4.96, p=0.002) and lactate dehydrogenase ≥450 IU/L (2.96, 1.60-5.45, p=0.001) were significantly associated with death. CONCLUSION: In this cohort of moderately and severely ill patients with COVID-19, severity of illness, underlying comorbidities and elevated levels of inflammatory markers were significantly associated with death. TRIAL REGISTRATION NUMBER: CTRI/2020/04/024775.

Analysis of a cholera toxin B subunit (CTB) and human mucin 1 (MUC1) conjugate protein in a MUC1-tolerant mouse model.

Since epithelial mucin 1 (MUC1) is associated with several adenocarcinomas at the mucosal sites, it is pertinent to test the efficacy of a mucosally targeted vaccine formulation. The B subunit of the Vibrio cholerae cholera toxin (CTB) has great potential to act as a mucosal carrier for subunit vaccines. In the present study we evaluated whether a MUC1 tandem repeat (TR) peptide chemically linked to CTB would break self-antigen tolerance in the transgenic MUC1-tolerant mouse model (MUC1.Tg) through oral or parenteral immunizations. We report that oral immunization with the CTB-MUC1 conjugate along with mucosal adjuvant, unmethylated CpG oligodeoxynucleotide (ODN) and interleukin-12 (IL-12) did not break self-antigen tolerance in MUC1.Tg mice, but induced a strong humoral response in wild-type C57BL/6 mice. However, self-antigen tolerance in the MUC1.Tg mouse model was broken after parenteral immunizations with different doses of the CTB-MUC1 conjugate protein and with the adjuvant CpG ODN co-delivered with CTB-MUC1. Importantly, mice immunized systemically with CpG ODN alone and with CTB-MUC1 exhibited decreased tumor burden when challenged with a mammary gland tumor cell line that expresses human MUC1.

Topological transition in a two-dimensional three-vector model: A non-Boltzmann Monte Carlo study.

Two-dimensional three-vector (d=2,n=3) lattice model of a liquid crystal (LC) system with order parameter space (R) described by the fundamental group Π_{1}(R)=Z_{2} was recently investigated based on non-Boltzmann Monte Carlo simulations. Its results indicated that the system did not undergo a topological transition condensing to a low temperature critical state as was reported earlier. Instead, a crossover to a nematic phase was observed, induced by the onset of a competing relevant length scale. This mechanism is further probed here by assigning a more restrictive R symmetry with Π_{1}(R)=Q (the discrete and non-Abelian group of quaternions), thus engaging the three spin degrees in the formation of point topological defects (disclinations). The results reported here indicate that such a choice of symmetry of the Hamiltonian with suitable model parameters leads to a defect-mediated transition to a low-temperature phase with topological order. It is characterized by a line of critical points with quasi-long-range order of its three spin degrees. The associated temperature-dependent power-law exponent decreases progressively and vanishes linearly as temperature tends to zero. The high-temperature disordered phase shows exponential spin correlations and their temperature-dependent lengths exhibit an essential singular divergence as the system approaches the topological transition point. Biaxial LC models have the required R symmetry owing to their tensor orientational orders and are suggested to serve as prototype examples to exhibit topological transition in (d=2,n=3) lattice models.

Breast-cancer-associated metastasis is significantly increased in a model of autoimmune arthritis.

INTRODUCTION: Sites of chronic inflammation are often associated with the establishment and growth of various malignancies including breast cancer. A common inflammatory condition in humans is autoimmune arthritis (AA) that causes inflammation and deformity of the joints. Other systemic effects associated with arthritis include increased cellular infiltration and inflammation of the lungs. Several studies have reported statistically significant risk ratios between AA and breast cancer. Despite this knowledge, available for a decade, it has never been questioned if the site of chronic inflammation linked to AA creates a milieu that attracts tumor cells to home and grow in the inflamed bones and lungs which are frequent sites of breast cancer metastasis. METHODS: To determine if chronic inflammation induced by autoimmune arthritis contributes to increased breast cancer-associated metastasis, we generated mammary gland tumors in SKG mice that were genetically prone to develop AA. Two breast cancer cell lines, one highly metastatic (4T1) and the other non-metastatic (TUBO) were used to generate the tumors in the mammary fat pad. Lung and bone metastasis and the associated inflammatory milieu were evaluated in the arthritic versus the non-arthritic mice. RESULTS: We report a three-fold increase in lung metastasis and a significant increase in the incidence of bone metastasis in the pro-arthritic and arthritic mice compared to non-arthritic control mice. We also report that the metastatic breast cancer cells augment the severity of arthritis resulting in a vicious cycle that increases both bone destruction and metastasis. Enhanced neutrophilic and granulocytic infiltration in lungs and bone of the pro-arthritic and arthritic mice and subsequent increase in circulating levels of proinflammatory cytokines, such as macrophage colony stimulating factor (M-CSF), interleukin-17 (IL-17), interleukin-6 (IL-6), vascular endothelial growth factor (VEGF), and tumor necrosis factor-alpha (TNF-alpha) may contribute to the increased metastasis. Treatment with anti-IL17 + celecoxib, an anti-inflammatory drug completely abrogated the development of metastasis and significantly reduced the primary tumor burden. CONCLUSIONS: The data clearly has important clinical implications for patients diagnosed with metastatic breast cancer, especially with regards to the prognosis and treatment options.

Natural fiber reinforced biodegradable staples: Novel approach for efficient wound closure.

Every year millions of lacerations and incisions taken place and require an effective methodology to manage the wound for a better life. The primary causes include mechanical trauma and surgical procedures. The rapid healing of the wound is critical to prevent further infection and reduction pain etc. Current options comprise of sutures, staplers, surgical strips and glues, again the intervention depends on the type of wound and the surgeon preference. The current wound closure techniques pose various potent limitations and confronting the problems to create a desired wound closure technique is necessary for faster and effective wound healing management. The surgical staplers are fast and easy to use wound closure devices, which approximates the edges of the wounds together by staples. The staples are mostly made up of metals like titanium and stainless steel. By modifying the existing stapling method using biodegradable staples that are expected to have good mechanical properties, not require removal procedure, minimized scarring and an overall acceleration in wound healing with minimal complications. Present, the paper focuses on the novel hypothesis on natural fiber reinforced biodegradable polymer staples as wound enclosures with high strength and degradability.

Complex free-energy landscapes in biaxial nematic liquid crystals and the role of repulsive interactions: A Wang-Landau study.

General quadratic Hamiltonian models, describing the interaction between liquid-crystal molecules (typically with D_{2h} symmetry), take into account couplings between their uniaxial and biaxial tensors. While the attractive contributions arising from interactions between similar tensors of the participating molecules provide for eventual condensation of the respective orders at suitably low temperatures, the role of cross coupling between unlike tensors is not fully appreciated. Our recent study with an advanced Monte Carlo technique (entropic sampling) showed clearly the increasing relevance of this cross term in determining the phase diagram (contravening in some regions of model parameter space), the predictions of mean-field theory, and standard Monte Carlo simulation results. In this context, we investigated the phase diagrams and the nature of the phases therein on two trajectories in the parameter space: one is a line in the interior region of biaxial stability believed to be representative of the real systems, and the second is the extensively investigated parabolic path resulting from the London dispersion approximation. In both cases, we find the destabilizing effect of increased cross-coupling interactions, which invariably result in the formation of local biaxial organizations inhomogeneously distributed. This manifests as a small, but unmistakable, contribution of biaxial order in the uniaxial phase. The free-energy profiles computed in the present study as a function of the two dominant order parameters indicate complex landscapes. On the one hand, these profiles account for the unusual thermal behavior of the biaxial order parameter under significant destabilizing influence from the cross terms. On the other, they also allude to the possibility that in real systems, these complexities might indeed be inhibiting the formation of a low-temperature biaxial order itself-perhaps reflecting the difficulties in their ready realization in the laboratory.

Surrogate in vitro activation of innate immunity synergizes with interleukin-7 to unleash rapid antigen-driven outgrowth of CD4+ and CD8+ human peripheral blood T-cells naturally recognizing MUC1, HER2/neu and other tumor-associated antigens.

Effective adoptive immunotherapy has proved elusive for many types of human cancer, often due to difficulties achieving robust expansion of natural tumor-specific T-cells from peripheral blood. We hypothesized that antigen-driven T-cell expansion might best be triggered in vitro by acute activation of innate immunity to mimic a life-threatening infection. Unfractionated peripheral blood mononuclear cells (PBMC) were subjected to a two-step culture, first synchronizing their exposure to exogenous antigens with aggressive surrogate activation of innate immunity, followed by γ-chain cytokine-modulated T-cell hyperexpansion. Step 1 exposure to GM-CSF plus paired Toll-like receptor agonists (resiquimod and LPS), stimulated abundant IL-12 and IL-23 secretion, as well as upregulated co-stimulatory molecules and CD11c expression within the myeloid (CD33+) subpopulation. Added synthetic long peptides (>20aa) derived from widely expressed oncoproteins (MUC1, HER2/neu and CMVpp65), were reliably presented to CD4+ T-cells and cross-presented to CD8+ T-cells. Both presentation and cross-presentation demonstrated proteasomal and Sec61 dependence that could bypass the endoplasmic reticulum. Step 2 exposure to exogenous IL-7 or IL-7+IL-2 produced selective and sustained expansion of both CD4+ and CD8+ peptide-specific T-cells with a predominant interferon-γ-producing T1-type, as well as the antigen-specific ability to lyse tumor targets. Other γ-chain cytokines and/or combinations were initially proliferogenic, but followed by a contractile phase not observed with IL-7 or IL-7+IL-2. Regulatory T-cells were minimally propagated under these culture conditions. This mechanistically rational culture sequence, effective even for unvaccinated donors, enables rapid preparation of T-cells recognizing tumor-associated antigens expressed by the majority of human cancers, including pancreatic cancers, breast cancers and glioblastomas.

Diagnosis of cystic echinococcosis in buffaloes by native 8 kDa antigen using latex agglutination test (LAT).

The dog tapeworm Echinococcusgranulosus is the causative agent of cystic hydatid disease in domestic/wild herbivores animals and man. Accurate immunodiagnosis of the infection requires highly specific and sensitive antigens. The aim of this study was to develop and evaluate immunoassays with principles of precipitation, agglutination for the identification of buffaloes infected with hydatid cyst which would allow the monitoring of animals from endemic areas and identifying infected animals prior to slaughter. The immunoassays were developed and validated using hydatid specific, non-cross reactive low molecular weight 8 kDa hydatid cyst fluid protein. Sera used for the assay validations were obtained from 200 buffaloes infected naturally with hydatid cyst and 200 non-infected buffaloes. The diagnostic sensitivity with latex agglutination test was 98.67 %. It should be useful for the conformation of hydatid cyst infected individual sheep.

Estimation of thyroglobulin in lymph node aspirates: Pilot experience from a tertiary referral cancer center.

BACKGROUND: Assessment of cervical lymph node involvement in patients with thyroid cancer either during preoperative surgical mapping or detection of recurrences during follow-up is a crucial step in the management of differentiated thyroid cancers (DTCs). In most patients, fine needle aspiration cytology (FNAC) confirms the presence of metastasis in lymph node. However, in cases of paucicellular lymph node aspirate or discordant sonogram and cytology results, thyroglobulin (Tg) measurement in the lymph node aspirate (FNA-Tg) is useful and a value >1 ng/ml is considered consistent with metastatic disease. CONTEXT: The addition of FNAC to the US improves the specificity, but 5-10% are nondiagnostic and 6-8% rate of false-negative results. Several studies have reported that the detection of Tg in FNA-needle washes improves the evaluation of suspicious lymph nodes in patients with DTC.Data from Indian centers on FNA-Tg are limited. AIMS: We piloted the utility of FNA-Tg in patients with sonographically suspicious cervical lymph node enlargement in the setting of suspicious thyroid nodule or in the follow-up of thyroid cancer. SETTINGS AND DESIGN: Prospective data collection. RESULTS: We measured Tg in 13 lymph node aspirates (12 patients, 10 females) among whom 4 patients had a total thyroidectomy and 1 had a hemithyroidectomy. Eight of the 13 lymph node aspirates had FNA-Tg values >150 ng/ml, all of them had unequivocal malignant cytology and four among them had proven metastatic DTC on surgical pathology. The median FNA-Tg of the patients with malignant cytology was 7550 ng/ml with a range of 162-30,000 ng/ml. Among the remaining 5 lymph node aspirate, 2 lymph nodes showed cytological features suggestive of reactive lymphadenitis (FNA-Tg <0.2 ng/ml) and were not operated, 1 had a high-grade malignancy consistent with anaplastic thyroid cancer (FNA-Tg <0.2 ng/ml), and 2 had nondiagnostic cytology (one had non-caseating granuloma on surgical pathology [FNA-Tg 1.3 ng/ml] and in the other patient [FNA-Tg <0.2 ng/ml] surgical intervention was deferred). CONCLUSIONS: FNA-Tg was concordant with positive cytology in all patients with DTC and may serve as a useful tool in patients with negative and nondiagnostic cytology to guide surgical management.

Aberrant Glycosylation of Anchor-Optimized MUC1 Peptides Can Enhance Antigen Binding Affinity and Reverse Tolerance to Cytotoxic T Lymphocytes.

Cancer vaccines have often failed to live up to their promise, although recent results with checkpoint inhibitors are reviving hopes that they will soon fulfill their promise. Although mutation-specific vaccines are under development, there is still high interest in an off-the-shelf vaccine to a ubiquitous antigen, such as MUC1, which is aberrantly expressed on most solid and many hematological tumors, including more than 90% of breast carcinomas. Clinical trials for MUC1 have shown variable success, likely because of immunological tolerance to a self-antigen and to poor immunogenicity of tandem repeat peptides. We hypothesized that MUC1 peptides could be optimized, relying on heteroclitic optimizations of potential anchor amino acids with and without tumor-specific glycosylation of the peptides. We have identified novel MUC1 class I peptides that bind to HLA-A*0201 molecules with significantly higher affinity and function than the native MUC1 peptides. These peptides elicited CTLs from normal donors, as well as breast cancer patients, which were highly effective in killing MUC1-expressing MCF-7 breast cancer cells. Each peptide elicited lytic responses in greater than 6/8 of normal individuals and 3/3 breast cancer patients. The CTLs generated against the glycosylated-anchor modified peptides cross reacted with the native MUC1 peptide, STAPPVHNV, suggesting these analog peptides may offer substantial improvement in the design of epitope-based vaccines.

MUC1 Vaccines, Comprised of Glycosylated or Non-Glycosylated Peptides or Tumor-Derived MUC1, Can Circumvent Immunoediting to Control Tumor Growth in MUC1 Transgenic Mice.

It remains challenging to produce decisive vaccines against MUC1, a tumor-associated antigen widely expressed by pancreas, breast and other tumors. Employing clinically relevant mouse models, we ruled out such causes as irreversible T-cell tolerance, inadequate avidity, and failure of T-cells to recognize aberrantly glycosylated tumor MUC1. Instead, every tested MUC1 preparation, even non-glycosylated synthetic 9mer peptides, induced interferon gamma-producing CD4(+) and CD8(+) T-cells that recognized glycosylated variants including tumor-associated MUC1. Vaccination with synthetic peptides conferred protection as long as vaccination was repeated post tumor challenge. Failure to revaccinate post challenge was associated with down-regulated tumor MUC1 and MHC molecules. Surprisingly, direct admixture of MUC1-expressing tumor with MUC1-hyperimmune T-cells could not prevent tumor outgrowth or MUC1 immunoediting, whereas ex vivo activation of the hyperimmune T-cells prior to tumor admixture rendered them curative. Therefore, surrogate T-cell preactivation outside the tumor bed, either in culture or by repetitive vaccination, can overcome tumor escape.