Use of pediatric thymus to humanize mice for HIV-1 mucosal transmission.

Humanized mice have been used to study human immunodeficiency virus type 1 (HIV-1) transmission, pathogenesis, and treatment. The ability of pediatric thymus tissue implanted either in the leg (Leg PedThy) or under the renal capsule (Renal PedThy) with allogeneic CD34+ hematopoietic cells (HSCs) in NSG mice was evaluated for reconstitution of human immune cells and for rectal transmission of HIV-1. These mice were compared to traditional BLT mice implanted with fetal liver and thymus under the renal capsule and mice injected only with HSCs. Renal PedThy mice had similar immune reconstitution in the blood, spleen and intestine as BLT mice, while Leg PedThy mice had transient detection of immune cells, particularly CD4+ T cells and macrophages, the target cells for HIV-1 infection. Rectal transmission and replication of HIV-1 was efficient in BLT mice but lower and more variable in Renal PedThy mice. HIV-1 was poorly transmitted in HSC mice and not transmitted in Leg PedThy mice, which correlated with the frequencies of target cells in the spleen and intestine. Humanization of NSG mice with pediatric thymus was successful when implanted under the kidney capsule, but led to less efficient HIV-1 rectal transmission and replication compared to BLT mice.

Cause-specific hazard regression estimation for modified Weibull distribution under a class of non-informative priors.

In time to event analysis, the situation of competing risks arises when the individual (or subject) may experience p mutually exclusive causes of death (failure), where cause-specific hazard function is of great importance in this framework. For instance, in malignancy-related death, colorectal cancer is one of the leading causes of the death in the world and death due to other causes considered as competing causes. We include prognostic variables in the model through parametric Cox proportional hazards model. Mostly, in literature exponential, Weibull, etc. distributions have been used for parametric modelling of cause-specific hazard function but they are incapable to accommodate non-monotone failure rate. Therefore, in this article, we consider a modified Weibull distribution which is capable to model survival data with non-monotonic behaviour of hazard rate. For estimating the cumulative cause-specific hazard function, we utilized maximum likelihood and Bayesian methods. A class of non-informative types of prior (uniform, Jeffrey's and half-t) is introduced for Bayes estimation under squared error (symmetric) as well as LINEX (asymmetric) loss functions. A simulation study is performed for a comprehensive comparison of Bayes and maximum likelihood estimators of cumulative cause-specific hazard function. Real data on colorectal cancer is used to demonstrate the proposed model.

CG Dinucleotide Removal in Bioluminescent and Fluorescent Reporters Improves HIV-1 Replication and Reporter Gene Expression for Dual Imaging in Humanized Mice.

Visualizing the transmission and dissemination of human immunodeficiency virus type 1 (HIV-1) in real time in humanized mouse models is a robust tool to investigate viral replication during treatments and in tissue reservoirs. However, the stability and expression of HIV-1 reporter genes are obstacles for long-term serial imaging in vivo. Two replication-competent CCR5-tropic HIV-1 reporter constructs were created that encode either nanoluciferase (nLuc) or a near-infrared fluorescent protein (iRFP) upstream of nef. HIV-1 reporter virus replication and reporter gene expression was measured in cell culture and in humanized mice. While reporter gene expression in vivo correlated initially with plasma viremia, expression decreased after 4 to 5 weeks despite high plasma viremia. The reporter genes were codon optimized to remove cytosine/guanine (CG) dinucleotides, and new CO-nLuc and CO-iRFP viruses were reconstructed. Removal of CG dinucleotides in HIV-1 reporter viruses improved replication in vitro and reporter expression in vivo and ex vivo. Both codon-optimized reporter viruses could be visualized during coinfection and in vivo reporter gene expression during treatment failure preceded detection of plasma viremia. While the dynamic range of CO-iRFP HIV-1 was lower than that of CO-nLuc HIV-1, both viruses could have utility in studying and visualizing HIV-1 infection in humanized mice. IMPORTANCE Animal models are important for studying HIV-1 pathogenesis and treatments. We developed two viruses each encoding a reporter gene that can be expressed in cells after infection. This study shows that HIV-1 infection can be visualized by noninvasive, whole-body imaging in mice with human immune cells over time by reporter expression. We improved reporter expression to reflect HIV-1 replication and showed that two viral variants can be tracked over time in the same animal and can predict failure of antiretroviral therapy to suppress virus.

Differential Activation of the Transcription Factor IRF1 Underlies the Distinct Immune Responses Elicited by Type I and Type III Interferons.

Type I and III interferons (IFNs) activate similar downstream signaling cascades, but unlike type I IFNs, type III IFNs (IFNλ) do not elicit strong inflammatory responses in vivo. Here, we examined the molecular mechanisms underlying this disparity. Type I and III IFNs displayed kinetic differences in expression of IFN-stimulated genes and proinflammatory responses, with type I IFNs preferentially stimulating expression of the transcription factor IRF1. Type III IFNs failed to induce IRF1 expression because of low IFNλ receptor abundance and insufficient STAT1 activation on epithelial cells and thus did not activate the IRF1 proinflammatory gene program. Rather, IFNλ stimulation preferentially induced factors implicated in tissue repair. Our findings suggest that IFN receptor compartmentalization and abundance confer a spatiotemporal division of labor where type III IFNs control viral spread at the site of the infection while restricting tissue damage; the transient induction of inflammatory responses by type I IFNs recruits immune effectors to promote protective immunity.

A high-throughput method of analyzing multiple plant defensive compounds in minimized sample mass.

PREMISE OF THE STUDY: Current methods for quantifying herbivore-induced alterations in plant biochemistry are often unusable by researchers due to practical constraints. We present a cost-effective, high-throughput protocol to quantify multiple biochemical responses from small plant tissue samples using spectrophotometric techniques. METHODS AND RESULTS: Using Solanum lycopersicum and Medicago polymorpha leaves pre- and post-herbivory, we demonstrate that our protocol quantifies common plant defense responses: peroxidase production, polyphenol oxidase production, reactive oxygen species production, total protein production, and trypsin-like protease inhibition activity. CONCLUSIONS: Current protocols can require 500 mg of tissue, but our assays detect activity in less than 10 mg. Our protocol takes two people 6 h to run any of the assays on 300 samples in triplicate, or all of the assays on 20 samples. Our protocol enables researchers to plan complex experiments that compare local versus systemic plant responses, quantify environmental and genetic variation, and measure population-level variation.

Endophytic Fungi-Alternative Sources of Cytotoxic Compounds: A Review.

Cancer is a major cause of death worldwide, with an increasing number of cases being reported annually. The elevated rate of mortality necessitates a global challenge to explore newer sources of anticancer drugs. Recent advancements in cancer treatment involve the discovery and development of new and improved chemotherapeutics derived from natural or synthetic sources. Natural sources offer the potential of finding new structural classes with unique bioactivities for cancer therapy. Endophytic fungi represent a rich source of bioactive metabolites that can be manipulated to produce desirable novel analogs for chemotherapy. This review offers a current and integrative account of clinically used anticancer drugs such as taxol, podophyllotoxin, camptothecin, and vinca alkaloids in terms of their mechanism of action, isolation from endophytic fungi and their characterization, yield obtained, and fungal strain improvement strategies. It also covers recent literature on endophytic fungal metabolites from terrestrial, mangrove, and marine sources as potential anticancer agents and emphasizes the findings for cytotoxic bioactive compounds tested against specific cancer cell lines.

Cholesterol homeostasis and cell proliferation by mitogenic homologs: insulin, benzo-α-pyrene and UV radiation.

Low-Density Lipoprotein (LDL) is known to promote the unregulated proliferation of cells that is progression of cancer. We aimed to investigate the effect of mitogens on the expression of cell cycle proteins, nuclear cholesterol and cell proliferation. We observed that insulin and benzo-α-pyrene (BaP) induced the expression of Low-Density Lipoprotein receptor (LDLR) on HepG2 cells, thereby enhancing the uptake of LDL. The internalized LDL increased the concentration of cholesterol in the cytoplasm and nucleus of the cell. At the same time, insulin and BaP also stimulated the expression of cell cycle proteins viz., Cyclin E and Cdk2, and thus induced more incorporation of Bromodeoxyuridine (BrdU) in cultured cells indicating increased DNA synthesis. Increased expression of cell cycle proteins and DNA synthesis are the indications of DNA replication and new cell synthesis. This suggests a link between the enhanced nuclear cholesterol concentration and new cell formation. On the other hand, UV irradiation with selectively given dose of cell death eventually decreases nuclear cholesterol concentration and LDLR expression. Reduced LDLR shows low functional activity. This, again, repeated the plausibility of the same link between intracellular cholesterol concentration and cell population. The biasness of adverse effect observed by UV irradiation has been compromised by inactivating LDLR with anti-LDLR antibody, resulting in similar effects on Cyclin E expression in the cultured cells. Hence, we concluded that in all the conditions, LDLR expression was found to be a translational event of its transcription factor, SREBP-2, by the induction of insulin, BaP and UV irradiation.

Differential Expressions of p53, p53R2, hRRM2 and PBR in Chronic Lymphocytic Leukemia: A Correlation with Intracellular Cholesterol.

Regulation of intracellular cholesterol homeostasis exists under balance between intracellular biosynthesis and uptake from extracellular origin by cell surface transport proteins. Expected role of cholesterol on either tumor suppressor gene and/or DNA synthesis has been aimed in the present study to explore intracellular cholesterol homeostasis in CLL subjects. Higher expressions of p53R2 (p53 dependent subunit of ribonucleotide reductase) and p53 were found in lymphocytes of chronic human lymphocytic leukemia as comparison to their normal counterparts. Inverse relation was found with p53 independent R2 subunit (in human hRRM2) of ribonucleotide reductase, which was found to be decreased from its control group. More expression of peripheral type benzodiazepine receptor, a cholesterol transporter, was noticed in isolated nuclear fraction with simultaneous increase of cholesterol concentration in cytoplasmic and nuclear compartments. A parallel increase of cholesterol in cell nucleus with increased p53R2 expression shows priority of the involvement of cholesterol in the process of cell replication.

Dermal Mucinosis in Chronic Sclerodermoid Graft-Versus-Host Disease.

Dermal mucinosis is characterized by the deposition of glycosaminoglycans (mucin), either focally or diffusely within the dermis. This may occur as a primary idiopathic disorder or secondary to several dermatoses, most notably lupus erythematous, scleroderma, and dermatomyositis. The authors present an unusual finding of dermal mucinosis in association with chronic sclerodermoid graft-versus-host disease.

Perforating lichen nitidus.

Transepidermal elimination and/or perforation of the epidermis is seen in a variety of conditions but has only rarely been reported in association with lichen nitidus. We describe a case of lichen nitidus with the unique finding of epidermal perforation, a feature that made the usual straightforward diagnosis of lichen nitidus much more difficult.

A rare case of melanoma with Touton-like giant cells: a potential diagnostic pitfall.

Although approximately 876,000 individuals in the United States currently have a diagnosis of melanoma, the Touton-like giant cell variant has been described only twice in the literature to date. In our case, a 70-year-old man with a history of sclerosing carcinoma on the scalp presented for evaluation of a new nodularity at the site of his previous surgery. On examination, a new complex pigmented lesion on the posterolateral scalp, adjacent to the recurrent sclerosing carcinoma, was noted. Biopsy of the pigmented lesion revealed an invasive melanoma with a Breslow depth of at least 2.78 mm. Microscopic sections showed a predominantly dermal-based tumor composed of sheets and nests of enlarged epithelioid cells. These cells contained oval-to-reniform nuclei with prominent nucleoli and an abundant amount of eosinophilic to vacuolated cytoplasm. Interestingly, numerous multinucleated melanocytes, some with a "Touton" appearance, were scattered throughout the lesion. The lesional cells demonstrated positivity to Mart-1 and HMB-45. Fortunately, the patient's sentinel lymph node biopsy was negative for micrometastases, and a subsequent Position Emission Tomography (PET) scan was unremarkable. Documentation of individual cases of this rare histologic variant of melanoma is necessary given the ability of this lesion to mimic benign histiocytic proliferations at scanning magnification.

Atypical fibrous histiocytoma arising in the perianal area: a case report and review of the literature.

Atypical fibrous histiocytoma (AFH) is an uncommon variant of cutaneous fibrous histiocytoma that can display histologic features associated with malignancy. Fewer than 150 cases have been reported in the literature. The majority of these lesions present on the trunk and extremities of middle-aged women. Genital lesions are rare, with one documented case of vulvar AFH and another case of scrotal AFH in the literature. We report an additional case of a 68-year-old woman who was diagnosed with an AFH in an unusual location, the perianal area. Histologically, the lesion was characterized by a nodular fairly well-circumscribed proliferation of large epithelioid macrophages with scattered lymphocytes and mast cells in the background. The atypical macrophages contained enlarged markedly pleomorphic nuclei with prominent nucleoli. Scattered multinucleated "monster cells" and atypical mitoses were observed throughout the lesion. Immunologically, the lesional cells were focally positive when stained with antibodies against CD163 and Factor XIIIa. They were negative for CD34, CD31, desmin, smooth muscle actin, CAM 5.2, keratin 5/6, S100, CD3, CD20, and CD30. The constellation of histologic and immunologic features was most consistent with an AFH. To our knowledge, this case is the first perianal presentation of AFH to date.

An unusual presentation and distribution of generalized eruptive syringomas.

A 19-year-old Caucasian man presented with numerous erythematous to flesh-colored papules that appeared in crops on his neck, axillae, buttocks, and lower back. The lesions started on his anterior neck at age 12. At 18 years, new crops of papules appeared on his axillae, back, and buttocks over several months. He reported pruritus in the lesions following exercise and perspiration. He denied any family history of similar lesions. His primary care physician treated him with topical triamcinolone 0.1% cream, which made the lesions smaller, less erythematous, and less pruritic; however, the papules never fully resolved. After discontinuation of the steroids, these erythematous pruritic papules gradually recurred in the same areas of his body. The patient denied any other medical complaints.

Reticulin and NM23 staining in the interpretation of lymph nodal nevus rests.

Melanocytic nevus rests in lymph nodes are a known diagnostic challenge, especially in patients with a history of melanoma. Reticulin and NM23 have been studied in this context. The pattern of reticulin staining in melanomas surrounds groups/nests of melanocytes but individual cells in benign nevi. NM23, a metastasis-suppressor gene, has an association with metastatic potential in melanomas and some carcinomas. Twenty-eight cases (14 cases of metastatic melanoma to lymph nodes and 14 cases of lymph node nevus rests, all confirmed with Melan-A staining) were stained with reticulin and NM23. The pattern of reticulin staining was reported as surrounding groups if staining was noted in approximately 5-10 melanocytes in greater than 50% of the lesion but was otherwise reported as surrounding individual melanocytes. Cytoplasmic staining was considered to represent reactivity for NM23. Reticulin staining around groups of melanocytes was identified in all 14 cases of metastatic melanoma. Regarding nodal nevus rest cases, 12 of 14 cases (86%) demonstrated staining around individual melanocytes, whereas in 2 cases, reticulin surrounded melanocytic groups. NM23 staining was equivocal in all cases. Reticulin staining reliably invests groups of melanocytes in cases of metastatic melanoma, whereas in nodal nevus rests, it predominantly surrounds individual melanocytes. NM23 demonstrated no discriminatory value in this analysis. In cases in which a collection of melanocytes is present within a lymph node, reticulin deposition around individual melanocytes supports a diagnosis of lymph nodal nevus rest.

BCR-ABL Translocation in Pediatric Acute Lymphoblastic Leukemia in Southern India.

Cytogenetics and polymerase chain reaction (PCR) based assays provide important information regarding biologically defined and prognostically relevant subgroups in acute leukemias. We utilized karyotyping and molecular analysis by reverse transcriptase PCR for the BCR-ABL translocation, in addition to morphological study, cytochemistry and immunophenotyping, to study 24 cases of pediatric acute lymphoblastic leukemia (ALL). Our objective was to determine the frequency of the BCRABL translocation in childhood ALL from southern India. Karyotyping showed one case of hyperdiploidy, one case of t (12; 21) translocation and one case of 46, XY-21+mar. The BCR-ABL translocation was found in 8.3% of these cases. One of these was a cryptic translocation, the karyotype being normal. BCR-ABL positivity in ALL is associated with aggressive disease and has been shown to be a poor prognostic factor, especially in children.

Ethnic differences in physiological cardiac adaptation to intense physical exercise in highly trained female athletes.

BACKGROUND: Ethnicity is an important determinant of cardiovascular adaptation in athletes. Studies in black male athletes reveal a higher prevalence of electric repolarization and left ventricular hypertrophy than observed in white males; these frequently overlap with those observed in cardiomyopathy and have important implications in the preparticipation cardiac screening era. There are no reports on cardiac adaptation in highly trained black females, who comprise an increasing population of elite competitors. METHODS AND RESULTS: Between 2004 and 2009, 240 nationally ranked black female athletes (mean age 21+/-4.6 years old) underwent 12-lead ECG and 2-dimensional echocardiography. The results were compared with 200 white female athletes of similar age and size participating in similar sports. Black athletes demonstrated greater left ventricular wall thickness (9.2+/-1.2 versus 8.6+/-1.2 mm, P<0.001) and left ventricular mass (187.2+/-42 versus 172.3+/-42 g, P=0.008) than white athletes. Eight black athletes (3%) exhibited a left ventricular wall thickness >11 mm (12 to 13 mm) compared with none of the white athletes. All athletes revealed normal indices of systolic and diastolic function. Black athletes exhibited a higher prevalence of T-wave inversions (14% versus 2%, P<0.001) and ST-segment elevation (11% versus 1%, P<0.001) than white athletes. Deep T-wave inversions (-0.2 mV) were observed only in black athletes and were confined to the anterior leads (V(1) through V(3)). CONCLUSIONS: Systematic physical exercise in black female athletes is associated with greater left ventricular hypertrophy and higher prevalence of repolarization changes than in white female athletes of similar age and size participating in identical sporting disciplines. However, a maximal left ventricular wall thickness >13 mm or deep T-wave inversions in the inferior and lateral leads are rare and warrant further investigation.

Genotoxic and antigenotoxic effects of Hemidesmus indicus R. Br. root extract in cultured lymphocytes.

AIM OF THE STUDY: The genotoxic and antigenotoxic potential of the ethanolic extract of Hemidesmus indicus roots were evaluated in cultured human lymphocytes using cisplatin as the positive mutagen. MATERIALS AND METHODS: Cytogenetic damage and cytotoxicity were determined in cells exposed to different doses of the extract, ranging from 2 to 32 microg/ml of culture medium, either alone or together with cisplatin. RESULTS: There was a significant reduction in cisplatin-induced frequencies of sister chromatid exchanges, chromosome aberrations and micronucleated binucleate cells at the lower concentrations of 4 and 8 microg/ml (P<0.05). However, the extract by itself reduced the proliferative rate index, mitotic index and cytokinesis-block proliferative index (P<0.05). Further, a significant increase in the percentage of chromosome aberrations was noticed at the higher concentrations. CONCLUSION: Hemidesmus indicus root extract possesses significant genoprotective effect at the lower concentrations although it is cytotoxic and probably genotoxic at higher doses.