7,12-Dimethylbenz[a]anthracene-Induced Malignancies in a Mouse Model of Menopause

Ovarian cancer has a high mortality rate because there are few symptoms in early disease development. The incidence of ovarian cancer increases in women after menopause. Understanding early events in this disease can best be achieved by using animal models. Therefore, the objective of this study was to develop and track the onset of ovarian tumorigenesis in mice mimicking characteristics of postmenopausal epithelial cancer in women. Female B6C3F1 mice (age, 28 d) received 4-vinylcyclohexene diepoxide (VCD, 160 mg/kg IV daily for 20 d) to cause ovarian failure. Four months after VCD treatment, via surgical intervention, each mouse received a single injection of 7,12-dimethylbenz[a]anthracene (DMBA) or vehicle control (sesame oil) under the bursa of the right ovary to cause ovarian neoplasms. The experimental groups were untreated controls (Con–Con), DMBA-treatment only (Con–DMBA), VCD treatment only (VCD–Con), and VCD+DMBA-treated (VCD+DMBA) mice. At 3, 5, 7, and 9 mo after DMBA injection, ovaries were collected for histologic and immunohistochemical evaluation. No tumors developed in Con–Con mice. All VCD-treated mice (with or without DMBA) exhibited ovarian failure. Mice that received both VCD and DMBA exhibited tumors at 3 mo (50%), 5 mo (14%), 7 mo (90%), and 9 mo (57%) after DMBA treatment; 31% of the tumors were epithelial in origin. Our findings confirm that inducing ovarian tumors in mice by chemical means is an effective method for studying early stages of tumor development that may be relevant to epithelial ovarian cancers that arise in postmenopausal women.Abbreviations: DMBA, 7,12-dimethylbenz[a]anthracene; VCD, 4-vinylcyclohexene diepoxideOvarian cancer, the most deadly female reproductive malignancy, has a high mortality rate because high-grade cancers are thought to metastasize early prior to the development of symptoms in early stages of disease.^4,27,28 The risk of contracting ovarian cancer over a lifetime is about 1 in 70, so it is a relatively rare cancer.²⁸ Although more than 20 types of ovarian malignancies exist, about 90% of human ovarian cancers are epithelial in origin.²⁸ Most cases are diagnosed at stages when the disease has metastasized outside the ovary, hindering efforts to treat or cure the disease. In addition, few reliable detection methods exist for early diagnosis of this disease. The incidence of ovarian cancer increases 8- to 10-fold among women in the peri- to postmenopausal period when compared with younger women.²⁸ The generation of animal models of ovarian cancer has been attempted for decades. These models have included whole-body irradiation,^5-7 chemical induction,^13,15,17,21 genetic manipulation,^18,25 and xenograph development.^9,23 It was observed as early as 1936 that the removal of all follicles from a mouse ovary was followed by the appearance of benign tubular adenomas in the residual ovarian tissue.^6,7,21,27 These adenomas appear to originate at the surface epithelium and proceed to invaginate and spread throughout the ovary.As women transition from peri- to postmenopause, circulating levels of estrogen and progesterone decrease, and the relative ratio of estrogens to androgens decreases in response to the decline of estrogen. In addition, gonadotropin (follicle stimulating hormone, luteinizing hormone) levels rise due to loss of negative feedback on the anterior pituitary and, thereafter, remain elevated.²⁶ One theory of ovarian carcinogenesis proposes that increased circulating gonadotropin levels after menopause contribute to the development of ovarian epithelial cancers by stimulating surface epithelium proliferation.¹⁸ Women who have undergone a natural progression to menopause have lost ovarian function but retain residual ovarian tissue. Therefore, because ovarian cancers in women arise more frequently after than before menopause, models developed in animals that have undergone ovarian failure and retain residual ovarian tissue likely most closely resemble the disease in postmenopausal women.Repeated daily dosing of mice with the ovotoxic chemical 4-vinylcyclohexene diepoxide (VCD) results in a gradual onset of ovarian failure.²⁴ Because VCD selectively targets primordial and primary follicles,²² larger follicles remain and develop toward ovulation.⁸ With the depletion of primordial and primary follicles, recruitment into the larger follicle pool eventually ceases, and a gradual onset of ovarian failure results. In VCD-treated mice, estrogen and progesterone concentrations decline and follicle-stimulating hormone levels rise after follicle depletion, similar to the scenario in postmenopausal women.¹⁹ A recent mouse model that combined virally induced changes in genes within the ovary and treatment with VCD resulted in ovarian failure along with induction of tumors characterized as undifferentiated tumors with mixed epithelial and stromal components along with some features of sex cord stromal tumors.¹⁸In a previous study, female Fisher 344 rats with VCD-induced ovarian failure developed ovarian tumors after treatment with 7, 12-dimethylbenz[a]anthracene (DMBA).¹¹ Specifically, 57% of the VCD+DMBA-treated rats developed ovarian tumors within 5 months after DMBA treatment. However, the tumors were all classified as Sertoli–Leydig cell type lesions, which are rare ovarian neoplasms in women and often much less aggressive than are their epithelial counterpart.^27,28 In another study,³ female B₆C₃F₁ mice were treated in the same way as in the Fisher 344 rat study.⁹ Similarly, all tumors that developed within 5 mo in treated mice (28%) were also Sertoli–Leydig cell type masses. Therefore, the present study was undertaken in B₆C₃F₁ mice to observe and classify DMBA-induced ovarian tumor development at later time points (7 and 9 mo after DMBA exposure) to determine whether epithelial tumors would develop and, if so, when.     

A Bayesian nonparametric model for zero‐inflated outcomes: Prediction,clustering, and causal estimation

Researchers are often interested in predicting outcomes, detecting distinct subgroups of their data, or estimating causal treatment effects. Pathological data distributions that exhibit skewness and zero‐inflation complicate these tasks—requiring highly flexible, data‐adaptive modeling. In this paper, we present a multipurpose Bayesian nonparametric model for continuous, zero‐inflated outcomes that simultaneously predicts structural zeros, captures skewness, and clusters patients with similar joint data distributions. The flexibility of our approach yields predictions that capture the joint data distribution better than commonly used zero‐inflated methods. Moreover, we demonstrate that our model can be coherently incorporated into a standardization procedure for computing causal effect estimates that are robust to such data pathologies. Uncertainty at all levels of this model flow through to the causal effect estimates of interest—allowing easy point estimation, interval estimation, and posterior predictive checks verifying positivity, a required causal identification assumption. Our simulation results show point estimates to have low bias and interval estimates to have close to nominal coverage under complicated data settings. Under simpler settings, these results hold while incurring lower efficiency loss than comparator methods. We use our proposed method to analyze zero‐inflated inpatient medical costs among endometrial cancer patients receiving either chemotherapy or radiation therapy in the SEER‐Medicare database.     

Cyclin K inhibits HIV-1 gene expression and replication by interfering with cyclin-dependent kinase 9 (CDK9)-cyclin T1 interaction in Nef-dependent manner

Human immunodeficiency virus-1 (HIV-1) exploits a number of host cellular factors for successful survival and propagation. The viral protein Nef plays an important role in HIV-1 pathogenesis by interacting with various cellular proteins. In the present work, we identified Cyclin K (CycK) as a novel Nef-interacting protein, and for the first time, we showed that CycK inhibits HIV-1 gene expression and replication in a Nef-dependent manner. The positive elongation factor b complex comprising cyclin-dependent kinase 9 (CDK9) and Cyclin T1 is a critical cellular complex required for viral gene expression and replication. Enhanced expression of CycK in the presence of Nef induced CycK-CDK9 binding, which prevented CDK9-Cyclin T1 complex formation and nuclear translocation of CDK9, resulting in inhibition of HIV-1 long terminal repeat-driven gene expression. Furthermore, this effect of CycK was not observed with Nef-deleted virus, indicating the importance of Nef in this phenomenon. Finally, silencing of CycK in HIV-1-infected cells resulted in increased translocation of CDK9 into the nucleus, leading to increased viral gene expression and replication. These data also suggest that endogenous CycK might act as an inhibitory factor for HIV-1 gene expression and replication in T-cells. Thus, our results clearly demonstrate that CycK utilizes HIV-1 Nef protein to displace CycT1 from the positive elongation factor b complex, resulting in inhibition of HIV-1 gene expression and replication.     

Synthesis, structure and theoretical studies of Hg(II)-NH carbene complex of annulated ligand pyridinyl[1,2-a]{2-pyridylimidazol}-3-ylidene hexaflurophosphate

Experimental and theoretical studies of a new class of novel annulated imidazolium scffords 2a, 2b, 2c and their Hg(II)-NHC complexes 3a, 3b and 3c have been synthesized and characterized by different spectroscopic techniques. Molecular structures of 3a and 3b have been determined by single crystal X-ray diffraction studies. Virtual linear geometry [C_carbene-Hg-C_carbene 176.56(17)°] around Hg of the complex 3b is observed where as in 3a and 3c its deviate a more. DFT calculations provide geometrical parameters in conformity with the experimental values in case of 3a. The molecules prefer syn configuration over the anti due to lower energy supported by theoretical studies. Attention has given paid for theoretical calculations and comparison of 3a and 3c to observe the steric effect of -CH₃ group in the scaffords where as in case of 3b the -CH₃ group remain innocent.     

Synthesis, characterization, electrochemical and magnetic studies on manganese(III) complexes involving bridging carboxylates

A series of carboxylate-bridged manganese(III) complexes derived from Schiff bases obtained by the condensation of salicylaldehyde or 5-bromo-salicylaldehyde and different types of diamine have been synthesized and characterized and, in the case of [Mn₂(L1)₂(μ-ClCH₂COO)](ClO₄) (1), the structure has been obtained by X-ray crystallography. The structure of 1 consists of two manganese atoms separated by 5.487(3) Å and bridged by a carboxylate anion. This dinuclear structural unit is linked by bridging phenoxy oxygens to adjacent dinuclear units to produce a one-dimensional chain. Cyclic voltammograms of all the compounds exhibit grossly similar features consisting of a reversible or quasi-reversible Mn^III/Mn^II reduction and a Mn^III/Mn^IV oxidation. It has been observed that bromo-substitution stabilizes the lower oxidation state in the Mn^III/Mn^II couple and destabilizes the higher oxidation state in the Mn^III/Mn^IV couple. Variable temperature magnetic susceptibility measurements of 1 show a weak antiferromagnetic interaction. The magnetic behavior is satisfactorily modeled by inclusion of zero-field splitting and an intermolecular interaction component.     

Structural basis of recognition of pathogen-associated molecular patterns and inhibition of proinflammatory cytokines by camel peptidoglycan recognition protein

Peptidoglycan recognition proteins (PGRPs) are involved in the recognition of pathogen-associated molecular patterns. The well known pathogen-associated molecular patterns include LPS from Gram-negative bacteria and lipoteichoic acid (LTA) from Gram-positive bacteria. In this work, the crystal structures of two complexes of the short form of camel PGRP (CPGRP-S) with LPS and LTA determined at 1.7- and 2.1-? resolutions, respectively, are reported. Both compounds were held firmly inside the complex formed with four CPGRP-S molecules designated A, B, C, and D. The binding cleft is located at the interface of molecules C and D, which is extendable to the interface of molecules A and C. The interface of molecules A and B is tightly packed, whereas that of molecules B and D forms a wide channel. The hydrophilic moieties of these compounds occupy a common region, whereas hydrophobic chains interact with distinct regions in the binding site. The binding studies showed that CPGRP-S binds to LPS and LTA with affinities of 1.6 × 10(-9) and 2.4 × 10(-8) M, respectively. The flow cytometric studies showed that both LPS- and LTA-induced expression of the proinflammatory cytokines TNF-α and IL-6 was inhibited by CPGRP-S. The results of animal studies using mouse models indicated that both LPS- and LTA-induced mortality rates decreased drastically when CPGRP-S was administered. The recognition of both LPS and LTA, their high binding affinities for CPGRP-S, the significant decrease in the production of LPS- and LTA-induced TNF-α and IL-6, and the drastic reduction in the mortality rates in mice by CPGRP-S indicate its useful properties as an antibiotic agent.     

Efflux transporters- and cytochrome P-450-mediated interactions between drugs of abuse and antiretrovirals

Multidrug regimens and corresponding drug interactions cause many adverse reactions and treatment failures. Drug efflux transporters: P-gp, MRP, BCRP in conjunction with metabolizing enzymes (CYPs) are major factors in such interactions. Most effective combination antiretrovirals (ARV) therapy includes a PI or a NNRTI or two NRTI. Coadministration of such ARV may induce efflux transporters and/or CYP3A4 resulting in sub-therapeutic blood levels and therapeutic failure due to reduced absorption and/or increased metabolism. A similar prognosis is true for ARV-compounds and drugs of abuse combinations. Morphine and nicotine enhance CYP3A4 and MDR1 expression in vitro. A 2.5 fold rise of cortisol metabolite was evident in smokers relative to nonsmokers. Altered functions of efflux transporters and CYPs in response to ARV and drugs of abuse may result in altered drug absorption and metabolism. Appropriate in vitro models can be employed to predict such interactions. Influence of genetic polymorphism, SNP and inter-individual variation in drug response has been discussed. Complexity underlying the relationship between efflux transporters and CYP makes it difficult to predict the outcome of HAART as such, particularly when HIV patients taking drugs of abuse do not adhere to HAART regimens. HIV(+) pregnant women on HAART medications, indulging in drugs of abuse, may develop higher viral load due to such interactions and lead to increase in mother to child transmission of HIV. A multidisciplinary approach with clear understanding of mechanism of interactions may allow proper selection of regimens so that desired therapeutic outcome of HAART can be reached without any side effects.