Stage of the Estrous Cycle Does Not Influence Myocardial Ischemia–Reperfusion Injury in Rats (Rattus norvegicus)

Even though cardiovascular disease is the leading cause of death for men and women, the vast majority of animal studies use male animals. Because female reproductive hormones have been associated with cardioprotective states, many investigators avoid using female animals because these hormones are cyclical and may introduce experimental variability. In addition, no studies have investigated the specific effects of the estrous cycle on cardiac ischemic injury. This study was conducted to determine whether the estrous cycle stage influences the susceptibility to ischemic injury in rat hearts. Estrous cycle stage was determined by using vaginal smear cytology, after which hearts underwent either in vivo (surgical) or ex vivo (isolated) ischemia–reperfusion injury. For in vivo studies, the left anterior coronary artery was ligated for 25 min of ischemia and subsequently released for 120 min of reperfusion. Infarct sizes were 42% ± 6%; 49% ± 4%; 40% ± 9%; 47% ± 9% of the zone-at-risk for rats in proestrus, estrus, metestrus, and diestrus, respectively. For ex vivo studies, isolated, perfused hearts underwent global ischemia and reperfusion for 25 and 120 min, respectively. Similar to our in vivo studies, the ex vivo rat model showed no significant differences in susceptibility to infarction or extent of cardiac arrhythmia according to estrous stage. To our knowledge, these studies provide the first direct evidence that the stage of estrous cycle does not significantly alter cardiac ischemia–reperfusion injury in rats.Abbreviations: VF, ventricular fibrillation; VT, ventricular tachycardiaCardiovascular disease remains the leading cause of morbidity and mortality throughout the industrialized world, with ischemic heart disease being a major manifestation of cardiovascular disease. Many investigators use animal models to advance our understanding of the etiology and mechanisms involved. Although ischemic heart disease is the leading cause of death for both men and women, the overwhelming majority of studies use male animals. Perhaps the most common reason for this practice is that physiologic fluctuations in female reproductive hormones such as estrogen may be a confounding variable, given the influence of female reproductive hormones on various organ systems.²⁵ Despite the assertion that cyclical variations in female reproductive hormones may confound experimental studies, few data are available that support estrous-cycle–dependent variations in susceptibility to ischemic heart injury.Epidemiologic studies suggest that, compared with men, women have lower cardiac mortality prior to undergoing menopause.⁴⁰ Consistent with human studies, experimental models in several species commonly show that the degree of cardiac injury in young female animals is lower than that in male counterparts.^7,9,21,22,42 Exogenous administration of estrogen has a clear effect in reducing injury,^14,15 but whether endogenous cyclical variations in female reproductive hormones affect cardiac injury is not known.Rats and mice are commonly used species to examine cardiac ischemia–reperfusion injury. Unlike humans, rodents do not undergo menstruation, during which the uterine endometrium sloughs off and is expelled through the vagina, but rather the uterine lining of rodents is reabsorbed during an estrous cycle.²⁴ The rat estrous cycle is typically 4 to 5 d in length and is defined by 4 separate stages: proestrus, estrus, metestrus, and diestrus. Proestrus is characterized by increasing levels of estrogen. At the end of proestrus, ovulation (signaled by luteinizing hormone) occurs and marks the beginning of the estrus cycle. During metestrus and diestrus, the uterine lining regenerates, and the cycle starts again.^24,33 These stages induce changes in the composition of the epithelium of the vagina and the presence of inflammatory cells, which can easily be detected by using vaginal cytology.^18,35We conducted the current study to determine whether estrous cycle stage influences the susceptibility to ischemia–reperfusion injury in the rat heart. Because the stage of the estrous cycle may influence cardiac injury either directly (via a direct effect of circulating hormones), or indirectly (by inducing changes that are intrinsic to the heart), we used both in vivo and ex vivo models of injury.     

Hydrogen sulfide mediates vasoactivity in an O2-dependent manner

Hydrogen sulfide (H(2)S) has recently been shown to have a signaling role in vascular cells. Similar to nitric oxide (NO), H(2)S is enzymatically produced by amino acid metabolism and can cause posttranslational modification of proteins, particularly at thiol residues. Molecular targets for H(2)S include ATP-sensitive K(+) channels, and H(2)S may interact with NO and heme proteins such as cyclooxygenase. It is well known that the reactions of NO in the vasculature are O(2) dependent, but this has not been addressed in most studies designed to elucidate the role of H(2)S in vascular function. This is important, since H(2)S reactions can be dramatically altered by the high concentrations of O(2) used in cell culture and organ bath experiments. To test the hypothesis that the effects of H(2)S on the vasculature are O(2) dependent, we have measured real-time levels of H(2)S and O(2) in respirometry and vessel tension experiments, as well as the associated vascular responses. A novel polarographic H(2)S sensor developed in our laboratory was used to measure H(2)S levels. Here we report that, in rat aorta, H(2)S concentrations that mediate rapid contraction at high O(2) levels cause rapid relaxation at lower physiological O(2) levels. At high O(2), the vasoconstrictive effect of H(2)S suggests that it may not be H(2)S per se but, rather, a putative vasoactive oxidation product that mediates constriction. These data are interpreted in terms of the potential for H(2)S to modulate vascular tone in vivo.     

Cytoplasmic localization of BAG-1 in leukoplakia and carcinoma of the tongue: correlation with p53 and c-erbB2 in carcinoma

BACKGROUND: The present study evaluated the clinical significance of BAG-1, an antiapoptotic protein, in leukoplakia and carcinoma of the tongue. METHODS: BAG-1 expression was evaluated by immunohistochemistry in paraffin-embedded tissues of leukoplakia (n=25) and carcinoma of the tongue (n=61). RESULTS: Cytoplasmic expression was predominantly seen in 80% and 70% of patients with leukoplakia and carcinoma, respectively. BAG-1 expression was found to be significantly lower in tobacco users than in non-tobacco users. BAG-1 expression in tobacco-using leukoplakia and carcinoma patients was compared by grouping the carcinoma patients according to lymph node status and disease stage. Carcinoma patients with tumor-positive lymph nodes had significantly lower BAG-1 expression than patients with negative lymph nodes and leukoplakia. Further, a trend towards an inverse correlation was observed with p53 and c-erbB2. In univariate and multivariate survival analysis, patient subgroups with 2+ or 3+ marker positivity (BAG-1 negativity, p53 and c-erbB2 positivity) had a reduced overall survival compared with patient subgroups with 1+ marker positivity or negativity. CONCLUSION: BAG-1 negativity in association with p53 and c-erbB2 positivity identified a subgroup of tongue cancer patients with an aggressive phenotype. Hence, an antiapoptotic protein, BAG-1, was found to be down-regulated in chewing-tobacco-mediated tongue carcinogenesis.     

Metabolic phenotyping of the Crohn's disease-like IBD etiopathology in the TNF(ΔARE/WT) mouse model

The underlying biochemical consequences of inflammatory bowel disease (IBD) on the systemic and gastrointestinal metabolism have not yet been fully elucidated but could help to better understand the disease pathogenesis and to identify tissue-specific markers associated with the different disease stages. Here, we applied a metabonomic approach to monitor metabolic events associated with the gradual development of Crohn's disease (CD)-like ileitis in the TNF(ΔARE/WT) mouse model. Metabolic profiles of different intestinal compartments from the age of 4 up to 24 weeks were generated by combining proton nuclear magnetic resonance ((1)H NMR) spectroscopy and liquid chromatography-mass spectrometry (LC-MS). From 8 weeks onward, mice developed CD similar to the immune and tissue-related phenotype of human CD with ileal involvement, including ileal histological abnormalities, reduced fat mass and body weight, as well as hallmarks of malabsorption with higher energy wasting. The metabonomic approach highlighted shifts in the intestinal lipid metabolism concomitant to the histological onset of inflammation. Moreover, the advanced disease status was characterized by a significantly altered metabolism of cholesterol, triglycerides, phospholipids, plasmalogens, and sphingomyelins in the inflamed tissue (ileum) and the adjacent intestinal parts (proximal colon). These results describe different biological processes associated with the disease onset, including modifications of the general cell membrane composition, alteration of energy homeostasis, and finally the generation of inflammatory lipid mediators. Taken together, this provides novel insights into IBD-related alterations of specific lipid-dependant processes during inflammatory states.     

ABL/BCR gene variant with two-step mechanism: Unusual localization and rare/novel chromosomal rearrangements in CML patients

Aims: Variant translocations involving 9q, 22q and at least one additional genomic locus occur in 5-10% of the patients with chronic myeloid leukemia (CML). The mechanisms for the formation of these variant translocations are not fully characterized. Here we report CML cases presenting a variant translocation indicating two-step mechanism with rare/novel chromosomal rearrangement. Methods: Karyotype analysis was performed on metaphases obtained through short-term cultures of bone marrow and blood. Detection of BCR-ABL fusion gene was performed using dual-color dual-fusion (D-FISH) and extra signal (ES) translocation probes. BAC-FISH was also carried out. Results: In Patient 1, the third partner chromosome was der(11)(p15) with a 2F2G1R signal pattern, which is an unusual signal pattern with the two-step mechanism. Patients 2 and 3 showed typical positive (2F1G1R) signal pattern. In Patient 2, both the chromosome 22s were involved in variant formation. The second fusion was observed below the BCR gene of the second homologue. In Patient 3 the third chromosome was der(13)(q14). The fourth patient showed a variant pattern with BCR/ABL-ES probe involving der(X)(q13) region. Conclusion: The presence of different rearrangements of both 9q34 and 22q11 regions highlights the genetic heterogeneity of this subgroup of CML. In each case with variants, further studies with FISH, BAC-FISH or more advanced technique such as microarray should be performed. Future studies should be performed to confirm the presence of true breakpoint hot spots and assess their implications in CML with variant Ph.     

Dendritic cell synthesis of C3 is required for full T cell activation and development of a Th1 phenotype

Previous studies have found that deficiency of complement component C3 is associated with reduced T cell responses in several disease models including viral infection, autoimmune disease, and transplantation. However, the underlying mechanism is unclear. In this study, we demonstrate that dendritic cells (DCs) are able to synthesize C3 and this synthesis is required for the capacity of DCs to stimulate alloreactive T cell responses in vitro and in vivo. Compared with C3-producing DCs, C3-nonproducing DCs exhibit reduced potency to stimulate an alloreactive T cell response, favor the polarization of CD4(+) T cells toward Th2 phenotype, and have regulatory T cell-driving capacity. In addition, priming mice with C3-deficient DCs compared with wild-type DCs led to delayed skin allograft rejection. Our findings that nonproduction of C3 by DCs significantly reduced T cell stimulation and impaired allograft rejection provide a potentially important explanation of how C3-deficient mice develop reduced T cell responses and of how C3-deficient donor kidney is protected from T cell-mediated graft rejection.     

Comparative study of serine-plasmalogens in human retina and optic nerve: identification of atypical species with odd carbon chains

The objective of this work was to detect and identify phosphatidylserine plasmalogen species in human ocular neurons represented by the retina and the optic nerve. Plasmalogens (vinyl-ether bearing phospholipids) are commonly found in the forms of phosphatidylcholine and phosphatidylethanolamine in numerous mammalian cell types, including the retina. Although their biological functions are unclear, the alteration of cellular plasmalogen content has been associated with several human disorders such as rhizomelic chondrodysplasia punctata Type 2 and primary open-angle glaucoma. By using liquid chromatography coupled to high-resolution and tandem mass spectrometry, we have identified for the first time several species of phosphatidylserine plasmalogens, including atypical forms having moieties with odd numbers of carbons and unsaturation in sn-2 position. Structural elucidation of the potential phosphatidylserine ether linked species was pursued by performing MS(3) experiments, and three fragments are proposed as marker ions to deduce which fatty acid is linked as ether or ester on the glycerol backbone. Interpretation of the fragmentation patterns based on this scheme enabled the assignment of structures to the m/z values, thereby identifying the phosphatidylserine plasmalogens.     

Synthesis and anti-leishmanial activity of 1-aryl-β-carboline derivatives against Leishmania donovani

β-carbolines from various natural and synthetic sources have been known to show diverse biological activities. As a part of our current ongoing project to search for potent natural product-derived anti-leishmanial compounds, we have synthesized a series of substituted 1-aryl-β-carboline derivatives. A total of 22 compounds were synthesized and tested in vitro against Leishmania donovani, out of which 6 compounds (4, 5, 10, 11, 19 and 22) showed notably more activity than the standard miltefosine (IC(50) 12.07±0.82 μM), with compound 4 being the most potent (IC(50) 2.16±0.26 μM).     

Coil embolization of intralobar pulmonary sequestration - an alternative to surgery: a case report

Background

Pulmonary sequestration is a congenital lung disease characterized by nonfunctioning pulmonary tissue that lacks normal communication with the bronchial tree and is supplied by a nonpulmonary systemic artery. Symptomatic bronchopulmonary sequestration is uncommon, seen more frequently in the pediatric population than in adults. It has traditionally been treated with surgical resection; however, a limited but growing number of cases have been treated with angiographic embolization. Given the inherent risks of cardiothoracic surgery, embolization of the anomalous vessel is an enticing alternative treatment. We present a case of a 56-year-old woman with known, symptomatic, intralobar pulmonary sequestration that was successfully treated with coil embolization.

Case presentation

A 56-year-old Pacific Islander woman with a history of chronic myeloid leukemia was admitted to the hospital with an episode of hemoptysis. Computed tomography of the chest demonstrated left lower lobe intralobar pulmonary sequestration fed by a large tortuous vessel branching off of the descending thoracic aorta. Surgical resection of the sequestration is the current standard treatment strategy of symptomatic intralobar pulmonary sequestration. The cardiothoracic surgeon noted that given the size and location of arterial blood supply, intervention would involve thoracotomy and lobectomy. The interventional radiologist offered embolization of the lesion as an alternative to surgery. Multiple coils, 6–13 mm in size, were used to embolize the sequestration. No considerable flow distal to the coils was noted postembolization.

Conclusions

Intralobar pulmonary sequestration is a rare condition that typically requires surgical management. This case demonstrates the efficacy of coil embolization as an alternative management strategy. To date, limited case reports of adults treated with endovascular embolization exist. Treatment of symptomatic pulmonary sequestration with embolization can be considered as an alternative to surgical resection.

     

LRH-1 drives colon cancer cell growth by repressing the expression of the CDKN1A gene in a p53-dependent manner

Liver receptor homologue 1 (LRH-1) is an orphan nuclear receptor that has been implicated in the progression of breast, pancreatic and colorectal cancer (CRC). To determine mechanisms underlying growth promotion by LRH-1 in CRC, we undertook global expression profiling following siRNA-mediated LRH-1 knockdown in HCT116 cells, which require LRH-1 for growth and in HT29 cells, in which LRH-1 does not regulate growth. Interestingly, expression of the cell cycle inhibitor p21 (CDKN1A) was regulated by LRH-1 in HCT116 cells. p21 regulation was not observed in HT29 cells, where p53 is mutated. p53 dependence for the regulation of p21 by LRH-1 was confirmed by p53 knockdown with siRNA, while LRH-1-regulation of p21 was not evident in HCT116 cells where p53 had been deleted. We demonstrate that LRH-1-mediated p21 regulation in HCT116 cells does not involve altered p53 protein or phosphorylation, and we show that LRH-1 inhibits p53 recruitment to the p21 promoter, likely through a mechanism involving chromatin remodelling. Our study suggests an important role for LRH-1 in the growth of CRC cells that retain wild-type p53.