Effects of vitamin D and estrogen receptor gene polymorphisms on the changes in lumbar bone mineral density with multiple pregnancies in Japanese women

BACKGROUND: Our aims were to follow the longitudinal changes in lumbar spine bone mineral density (BMD) with multiple pregnancies, and to study whether polymorphisms in the vitamin D receptor (VDR) and estrogen receptor (ER) genes may influence the results. METHODS: We repeatedly measured the BMD of the lumbar spine (L2-L4) of 133 women who had undergone two successive pregnancies and 73 non-pregnant controls, and analysed the restriction fragment length polymorphisms using restriction endonucleases TaqI, ApaI and FokI for the VDR gene, and PvuII and XbaI for the ER gene. RESULTS: Cases and controls had no significant differences in the longitudinal BMD changes. The mean percentage change in lumbar BMD (DeltaBMD%) of the women with the XX/Xx genotype was significantly lower than that of the women with the xx genotype after adjusting for age at each delivery, BMD of the first scan, and interval between the scans (0.2 +/- 3.3 versus 2.0 +/- 4.2%; P=0.030, analysis of covariance). Multiple regression analyses to evaluate the contribution of the XbaI polymorphism of the ER gene on DeltaBMD% showed that the percentage decrease in BMD was greater for women lacking the XbaI restriction site (adjusted R2=0.188, P<0.0001). CONCLUSIONS: The present study suggests that the DeltaBMD% was significantly influenced by the XbaI polymorphism of the ER gene.     

Hypoxia-induced renal epithelial cell death through caspase-dependent pathway: role of Bcl-2, Bcl-xL and Bax in tubular injury

Although injury of epithelial cells has been reported to be responsible for renal disease such as acute renal failure, its molecular mechanisms are largely unknown. As hypoxia has been postulated as the initial trigger of epithelial injury, we studied the molecular mechanisms of apoptosis induced by hypoxia in human renal epithelial cells. Severe hypoxia caused epithelial cell death, accompanied by a significant increase in LDH release (p<0.01). In addition, hypoxic treatment of epithelial cells resulted in a significant increase in apoptotic cells as assessed by cell morphology (p<0.01). The apoptotic change in epithelial cells under hypoxic condition was also confirmed by a significant increase in caspase-3-like activity and release of cytochrome c (p<0.01). The decrease in epithelial cell number was completely abolished by addition of a wide-spectrum caspase inhibitor, Z-VAD, rather than Z-DEVD, a specific caspase-3 inhibitor (p<0.01). Thus, we further studied the molecular mechanisms of apoptosis induced by hypoxia. Anti-apoptotic factors, Bcl-2 and Bcl-xL, were significantly decreased in epithelial cells under a hypoxic condition as assessed by Western blotting (p<0.01). In contrast, hypoxia did not alter their location. Of particular importance, translocation of a proapoptotic factor, Bax, from the cytoplasm to the mitochondrial membrane was observed in response to hypoxia, whereas total Bax protein was not changed by hypoxia. Overall, this study demonstrated that hypoxia caused epithelial cell death induced by caspase-3-like activity-dependent apoptosis. The pro-apoptotic mechanisms of hypoxia in epithelial cells largely depend on a significant decrease in Bcl-2 and Bcl-xL. In addition, the present results demonstrate that translocation of Bax from the cytosol to the mitochondrial membrane occurred under hypoxia, thereby leading to pathological tissue destruction.     

Evidence for the presence of immunoglobulin E antibodies specific to the cell wall phosphomannoproteins of Candida albicans in patients with allergies.

To determine the major antigenic component of Candida albicans against immunoglobulin E (IgE) antibodies in the sera of patients with allergies who were positive for IgE antibodies to C. albicans crude antigen in a CAP system, phosphomannoproteins (CAMP/A or CAMP/B for serotype A or B strain, respectively) and their acid-stable portions (CAMP-S/A or CAMP-S/B) were isolated from beta-mercaptoethanol (2-ME) extracts of C. albicans cells of serotypes A and B, and IgE antibodies against these components were compared with those against protein complex and enolase (CAE) fractions isolated from C. albicans cells. The dot blot test, which was used to detect IgE antibodies to the C. albicans antigens, showed that IgE antibodies to the 2-ME extract and phosphomannoprotein fractions were present in the sera of 98.0% (2-ME extract), 96.8% (CAMP/A), 93.2% (CAMP-S/A), 97.2% (CAMP/B), and 81.5% (CAMP-S/B) of the patients, whereas IgE antibodies to the protein complex and CAE fractions were found in the sera of 73.6 and 48.8% of the patients, respectively. The extent of IgE binding to the 2-ME extract and phosphomannoproteins was well correlated with the fluorescence intensities estimated with the CAP system. Furthermore, the results obtained from the inhibition experiment with the CAP system indicated that the binding of IgE antibodies to Candida antigens is strongly inhibited by the phosphomannoprotein fraction and is an indication that the serum of the patients contained IgE antibodies specific to the cell wall phosphomannoproteins of C. albicans. Finally, an initial chemical analysis indicated that the epitopes for IgE antibodies on the phosphomannoproteins is a carbohydrate portion, since the ability of CAMP/A to inhibit the binding of IgE antibodies to the homologous CAMP/A was destroyed after oxidation by sodium periodate but not after digestion with proteinase K.     

Lymphatic invasion according to D2-40 immunostaining is a strong predictor of nodal metastasis in superficial squamous cell carcinoma of the esophagus: algorithm for risk of nodal metastasis based on lymphatic invasion

In squamous cell carcinoma (SCC) of the esophagus, D2-40 immunostaining has recently been used to detect lymphatic invasion, but invasion detected using D2-40 immunostaining for a predictor of nodal metastasis was controversial. Therefore, the usefulness of detecting lymphatic invasion by D2-40 immunostaining as a predictor of nodal metastasis was examined in superficial (mucosal and submucosal) SCC of the esophagus. A total of 115 superficial SCC of the esophagus were examined on immunohistochemistry using D2-40. It was found that lymphatic invasion demonstrated on D2-40 immunostaining was mainly detected in the lamina propria mucosa. Lymphatic invasion was found in 37 cases and the invasion detected in the entire tumor tissue was statistically correlated with nodal metastasis. Based on the lymphatic invasion according to D2-40 immunostaining, an algorithm was devised for the risk (low, intermediate and high) of nodal metastases in superficial SCC in the esophagus. In conclusion, the detection of lymphatic invasion on D2-40 immunostaining in tumor tissue is a strong predictor for nodal metastasis in superficial SCC of the esophagus. Lymphatic invasion was found mainly in the lamia propria mucosa, thus the devised algorithm is useful for determining the optimal treatment strategy after endoscopic mucosal resection for esophageal SCC.     

mRNA expression changes of slit proteins following peripheral nerve injury in the rat model

Slit family of proteins is one of the repulsive axonal guidance cues, and it also plays an important role in neuronal migration and branching through the interaction with roundabout receptors. The function and role of Slit family proteins during peripheral nerve regeneration are still unknown. We examined the expressions of Slits 1-3 mRNAs in the facial nerve nuclei after facial nerve transection by in situ hybridization, using Sprague-Dawley rats. Slit 1 mRNA was weakly expressed in the facial motoneurons, and its expression increased from day 5 to day 28 after transection, with the peak on day 14 after axotomy. Slits 2 and 3 mRNAs were expressed in the motoneurons of the facial nerve before injury, but the expression of Slit 2 mRNA was down-regulated from day 1 to day 7 after axotomy, with the peak on the first day after injury. Slit 3 mRNA expression in the axotomized side remained unchanged throughout the examination period. Slits 1 and 2 mRNA expression returned to the normal level on day 56 postoperatively. The difference in expression pattern of Slit family mRNA in the neurons during peripheral nerve regeneration suggests that it plays a different role in axonal regeneration after axotomy of peripheral nerves.     

Evaluation of left ventricular mechanical work and energetics of normal hearts in SERCA2a transgenic rats

Cardiac sarcoplasmic reticulum (SR) Ca(2+)-ATPase (SERCA2a) is responsible for most of the Ca(2+) removal during diastole and a larger Ca(2+) handling energy consumer in excitation-contraction (E-C) coupling. To understand the cardiac performance under long-term SERCA2a overexpression conditions, we established SERCA2a transgenic (TG) Wistar rats to analyze cardiac mechanical work and energetics in normal hearts during pacing at 300 beats/min. SERCA2a protein expression was increased in TGI and TGII rats (F2 and F3 of the same father and different mothers). Mean left ventricular (LV) end-systolic pressure (ESP) and systolic pressure-volume area (PVA; a total mechanical energy per beat) at midrange LV volume (mLVV) were significantly larger in TGI rats and were unchanged in TGII rats, compared to those in non-TG [wildtype (WT)] littermates. Mean myocardial oxygen consumption per minute for E-C coupling was significantly increased, and the mean slope of myocardial oxygen consumption per beat (VO(2))-PVA (systolic PVA) linear relation was smaller, but the overall O(2) cost of LV contractility for Ca(2+) is unchanged in all TG rats. Mean Ca(2+) concentration exerting maximal ESP(mLVV) in TGII rats was significantly higher than that in WT rats. The Ca(2+) overloading protocol did not elicit mitochondrial swelling in TGII rats. Tolerance to higher Ca(2+) concentrations may support the possibility for enhanced SERCA2a activity in TGII rats. In conclusion, long-term SERCA2a overexpression enhanced or maintained LV mechanics, improved contractile efficiency under higher energy expenditure for Ca(2+) handling, and improved Ca(2+) tolerance, but it did not change the overall O(2) cost of LV contractility for Ca(2+) in normal hearts of TG rats.     

HIF-1 in T cells ameliorated dextran sodium sulfate-induced murine colitis

HIF-1 is active in hypoxia, such as inflamed mucosa, and HIF-1 in epithelium has been reported to control inflamed mucosa in IBD models. Although T cells play an important role for pathogenesis of IBD, the function of HIF-1 in T cells remains to be elucidated. We aimed to clarify the function of HIF-1 in T cells in IBD with focus on the balance between Treg and Teff. Double immunohistochemistry of colonic mucosa in IBD patients showed that HIF-1 was expressed in T cells infiltrating the inflamed mucosa, suggesting that HIF-1 in T cells is involved in the pathogenesis. DSS administration to T cell-specific HIF-1α KO mice showed more severe colonic inflammation than control mice with the up-regulation of Th1 and Th17. Hypoxic stimulation in vitro increased Treg activation in WT T cells but not in HIF-1-deleted T cells. In contrast, hypoxic stimulation increased Th17 activation, and the degree was higher in HIF-1-deleted cells than in control cells. These results show that hypoxia controls intestinal inflammation by regulating cytokine balance in a HIF-1-dependent manner, suggesting that strengthening HIF-1 induction in T cells at the sites of inflammation might be a therapeutic strategy for IBD regulation.     

fMRI of patients with social anxiety disorder during a social situation task

Previous functional neuroimaging studies found that the amygdala and other limbic regions may play a substantial role in social anxiety disorder (SAD). However, more widely distributed large-scale brain systems may be involved in cognitive processing in SAD patients when confronted with social situations. We employed functional MRI (fMRI) to investigate local brain activation of patients with SAD (n=6) and healthy controls (HC, n=9) during cognitive work. During fMRI scanning, subjects performed a social situation task using a block design paradigm in which the task and control trials were performed by turn. The patients with SAD showed higher anxiety levels during scanning in all social situations. The HC group showed greater common activation in the posterior cingulate cortex (PCC), cuneus, occipital gyrus, and cerebellum. Although the patients with SAD showed activation patterns similar to that of the HC group, they showed comparatively significant decreased activation in the left cerebellum, left precuneus, and bilateral PCC. The present study demonstrates that SAD may involve dysfunction of a broad neuronal network including the limbic system, parieto-posterior cortex and cerebellum. The findings contribute to previous findings that revealed abnormal activities of emotion-related regions including the amygdala and insular cortex during facial perception in SAD.     

Microsatellite-based genotyping of Candida albicans isolated from patients with superficial candidiasis

This study aimed to examine the genotype distribution of Candida albicans and the major genotypes involved in superficial candidiasis. The genotypes of C. albicans isolated from the infection sites of patients with superficial candidiasis (referred to as infection isolates) were analyzed by fragment analysis using 4 microsatellite markers (HIS3, CDC3, CAI and CAIII). Genotypes of the infection isolates were compared with those of C. albicans isolated from oral mucosa of non-candidiasis patients (referred to as oral isolates). Isolates of C. albicans showed 4 major genotypes for HIS3/CAI (" a " for 148 : 148 / 23 : 23," b " for 148 : 160 / 33 : 41," c " for 148 : 164 / 32 : 41 and " d " for 152 : 152 / 18 : 27). The genotypes " a "," b " and " d " were commonly found in oral (4.7, 8.8 and 7.6%, respectively) and infection (6.6, 9.2 and 15.4%, respectively) isolates. No isolates of genotype " c " were isolated from infection sites. The genotype " a " was found in the isolates from patients with genitalia candidiasis. Genotyping of multiple isolates from an individual patient showed that C. albicans from infection sites was genetically homogenous as compared with that of oral isolates, even in the same patient with candidiasis.     

B-cell activation in the mesenteric lymph nodes of resistant BALB/c mice infected with the murine nematode parasite Trichuris muris

Immune responses in resistant BALB/c mice infected with the murine nematode parasite Trichuris muris were examined. Following the establishment of infection, worm burdens of T. muris were expelled by BALB/c mice by day 21 postinfection (p.i.). Specific immunoglobulin G₁ (IgG₁) antibodies to T. muris excretory/secretory (E/S) antigens were detected in sera from infected mice, though specific IgG_2a antibodies were not observed during infection. Ig-producing cells increased in the mesenteric lymph nodes (MLN) of infected mice on days 7, 14, and 21 p.i., with the greatest increase in numbers of IgG- and IgA-producing cells occurring on day 14. Marked increases in the relative percentages of B220⁺ and surface Ig⁺ (sIg⁺) cells were observed in the MLN of infected mice on days 14 and 21 p.i. Furthermore, cellular expansion of the MLN in infected mice resulted in an increase in the absolute numbers of B220⁺ and sIg⁺ cells. The levels of interleukin 2 (IL-2), IL-4, and interferon-γ (IFN-γ) detected in the supernatants from concanavalin A-stimulated MLN cells of infected mice were higher than those found in normal mice. Consequently, the expulsion of T. muris in resistant BALB/c mice was concomitant with cytokine production and B-cell activation in the MLN of infected mice. These results suggest the involvement of B-cell responses in protective immunity to T. muris infection. Received: 12 May 1998 / Accepted: 5 August 1998