Plasma kisspeptin levels in lactational amenorrhea.

The kisspeptin is a neuropeptide to play physiological roles in regulating gonadotropin-releasing hormone secretion in the hypothalamus. In human plasma, the kisspeptin concentration is measured, but gonadotropin-releasing hormone is not. This study aims to understand the physiological roles of the circulating kisspeptin in lactational amenorrhea in humans because prolactin reduces the kisspeptin expression and luteinizing hormone secretion resulting in anovulations in rodent brains. Plasma kisspeptin levels were measured in 11 subjects in lactational amenorrhea and in four cases with pathological amenorrhea by different etiologies for comparison using the enzyme immunoassay specific for human kisspeptin. The plasma kisspeptin levels in the 11 women with lactational amenorrhea were 15.2 ± 2.5 fmol/mL (mean ± SD) which were not significantly different as compared with 16.5 ± 4.8 fmol/mL (mean ± SD) in four age-matched women with menstrual cycles as we reported previously. In the four cases with pathological amenorrhea, their plasma kisspeptin levels were from 5.8 to 13.7 fmol/mL. This study demonstrated that the plasma kisspeptin levels were not totally reduced in lactational or pathological amenorrhea. These results suggest the physiological roles of the circulating kisspeptin are different from the role in the brain.

Inflammation-Related Tumor Progression in Murine Fibrosarcoma Exhibited Over-expression of Sex-determining Region Y-box 2 (Sox2) Compared to Parental Regressor Cells.

BACKGROUND/AIM: Tumor progression is one of the most serious issues to overcome cancer disease. As a model of inflammation-induced tumor progression, we used the regressive murine fibrosarcoma cell clone QR-32 and the progressive malignant clone QRsP-11, that was derived from QR-32. Heat shock protein beta-1 (Hspb1) is a molecular chaperone. Hspb1 plays roles in not only cell protection but also chemo-resistance, tumorigenicity and protection from apoptosis. In a recent study, we showed that Hspb1 was up-regulated in QRsP-11 compared to QR-32. MATERIALS AND METHODS: We compared the expression levels of Hspb1, Hsf1 and Sox2 in QR-32 and QRsP-11 cells by means of western blotting. RESULTS: Hsf1, a transcription factor for Hspb1 was not increased in QRsP-11. Sex determining region Y-box 2 (Sox2) is a transcription factor, reported to interact with Hspb1. Sox2 was up-regulated in QRsP-11 compared to QR-32. CONCLUSION: These results suggest that Sox2-Hspb1 signaling is a possible pathway responsible to tumor progression of QRsP-11.

Serum levels of IgG and IgG4 in Hashimoto thyroiditis.

Although IgG4-related disease is characterized by extensive infiltration of IgG4-positive plasma cells and lymphocytes of various organs, the details of this systemic disease are still unclear. We screened serum total IgG levels in the patients with Hashimoto thyroiditis (HT) to illustrate the prevalence of IgG4-related thyroiditis in HT. Twenty-four of 94 patients with HT (25.5%) had elevated serum IgG levels and their serum IgG4 was measured. Five of the 24 cases had more than 135 mg/dL of IgG4, which is the serum criterion of IgG4-related disease. One was a female patient who was initially treated as Graves' disease and rapidly developed a firm goiter and hypothyroidism. The biopsy of her thyroid gland revealed that follicular cells were atrophic with squamous metaplasia, replaced with fibrosis, which was compatible with the fibrous variant of HT. Immunohistochemical examination revealed diffuse infiltration of IgG4-positive plasma cells, and the serum IgG4 level was 179 mg/dL. The levels of IgG and IgG4 were positively correlated with the titers of anti-thyroglobulin antibody or anti-thyroid peroxidase antibody. In conclusion, at least a small portion of patients with HT with high titers of anti-thyroid antibodies may overlap the IgG4-related thyroiditis.

Integrated network systems and evolutionary developmental endocrinology.

Endocrine system has been considered to be a linear one, but the 'real world endocrine system' is a complex system, which is difficult to investigate using conventional strategies, such as single nucleotide polymorphism, genome-wide analysis, or gene targeting in animals. Here we propose a new strategy to comprehend the endocrine system as a complex network system. We introduced several novel concepts, such as complex system, network analysis, systems biology and evolutionary medicine, into the comprehension of endocrine system as a whole complex network system. This system is considered to be a scale-free network with key molecules such as acetyl CoA, NAD or ATP as 'hubs'. This system is robust against simple mutations, but various complex diseases may attack hubs. The system is also 'fractals', since there exist similar network systems among cells, proteins, and transcription factors in the lower levels, and there are similar ones among disease and social network in the higher levels. We propose to call this model 'Integrated Network Systems and Evolutionary DEvelopmental ENdocrinology (INS-EDEN)'. This novel framework will facilitate us to develop a new approach for understanding and treatment of various complex diseases related to endocrinology, and identify a unified theory of complex diseases.

In vitro differentiation of Runx3-/- p53-/- gastric epithelial cells into intestinal type cells.

We have reported that a lack of RUNX3 function is causally associated with gastric carcinogenesis. We have also presented evidence that loss of Runx3 may be related to the genesis of intestinal metaplasia because expression of RUNX3 is reduced in some intestinal metaplasias, and some Runx3(-/-)p53(-/-) gastric epithelial cells differentiate into intestinal type cells in vivo. Recently several reports have indicated that blood cells play important roles in the gastric carcinogenesis. In the present study, we therefore examined whether Runx3(-/-)p53(-/-) gastric epithelial cells differentiate autonomously into intestinal type cells, or whether the presence of other cells is necessary for the differentiation in vitro. When Runx3(-/-)p53(-/-) gastric epithelial cells were cultured with collagen gels, they did not exhibit any morphogenesis and differentiated poorly. When cultured with fetal mouse gastric mesenchymes, the cells formed glandular structures and differentiated into surface mucous cells, but differentiation of intestinal type cells was never observed. When cultured with Matrigel, the cells formed glandular structures, and some cells differentiated into intestinal type cells in vitro. Reverse transcription-polymerase chain reaction analysis showed that the cells expressed stomach-specific genes, and their levels decreased gradually during the culture. The cells expressed some intestine-specific genes weakly at the start of culture, and their levels were increased with time in culture. We therefore conclude that Runx3(-/-)p53(-/-) gastric epithelial cells differentiate into intestinal type cells in combination with Matrigel in the absence of other cell types. Extracellular matrix, not blood cells, may play a role in the genesis of intestinal metaplasia.

Reversible posterior leukoencephalopathy in a patient with Wegener granulomatosis.

A 14-year-old girl with rapidly progressive glomerulonephritis was transferred to our hospital because of acute renal failure. A diagnosis of Wegener granulomatosis was made according to the symptom triad of a renal biopsy demonstrating crescentic glomerulonephritis, severe sinusitis, and serological findings of raised proteinase 3 anti-neutrophil cytoplasmic antibody level. In spite of combination therapy with methylprednisolone, cyclophosphamide, and plasma exchange, her renal function gradually deteriorated. Thereafter, she suffered a severe headache and generalized seizures. Brain computed tomography (CT) scan revealed bilateral low-density areas in the parieto-occipital lobes. Magnetic resonance imaging (MRI) disclosed a high-intensity area on T2-weighted images and a low-signal intensity area on T1-weighted images in the same lesion. Follow-up brain CT scan 3 weeks and MRI 2 months after the first studies showed complete resolution of the abnormal lesions, which indicated reversible posterior leukoencephalopathy syndrome. In addition to renal failure, hypertension, and cyclophoshamide, the primary disease may have played a role in the development of this uncommon syndrome in our patient.

Role of prostaglandin E receptor EP1 subtype in the development of renal injury in genetically hypertensive rats.

One of the major causes of end-stage renal diseases is hypertensive renal disease, in which enhanced renal prostaglandin (PG) E2 production has been shown. PGE2, a major arachidonic acid metabolite produced in the kidney, acts on 4 receptor subtypes, EP1 through EP4, but the pathophysiological importance of the PGE2/EP subtypes in the development of hypertensive renal injury remains to be elucidated. In this study, we investigated whether an orally active EP1-selective antagonist (EP1A) prevents the progression of renal damage in stroke-prone spontaneously hypertensive rats (SHRSP), a model of human malignant hypertension. Ten-week-old SHRSP, with established hypertension but with minimal renal damage, were given EP1A or vehicle for 5 weeks. After the treatment period, vehicle-treated SHRSP showed prominent proliferative lesions in arterioles, characterized by decreased alpha-smooth muscle actin expression in multilayered vascular smooth muscle cells. Upregulation of transforming growth factor-beta expression and tubulointerstitial fibrosis were also observed in vehicle-treated SHRSP. All these changes were dramatically attenuated in EP1A-treated SHRSP. Moreover, EP1A treatment significantly inhibited both increase in urinary protein excretion and decrease in creatinine clearance but had little effect on systemic blood pressure. These findings indicate that the PGE2/EP1 signaling pathway plays a crucial role in the development of renal injury in SHRSP. This study opens a novel therapeutic potential of selective blockade of EP1 for the treatment of hypertensive renal disease.

Roles of connective tissue growth factor and prostanoids in early streptozotocin-induced diabetic rat kidney: the effect of aspirin treatment.

BACKGROUND: Connective tissue growth factor (CTGF) is a cysteine-rich growth factor induced by transforming growth factor-beta (TGF-beta) and is thought to play a critical role in TGF-beta-stimulated extracellular matrix accumulation. To explore its involvement in early diabetic nephropathy, we investigated the time course of CTGF gene expression and its regulation in streptozotocin (STZ)-induced diabetic rat kidney. METHODS: Northern blot analysis for CTGF, TGF-beta, and fibronectin expression was performed in the glomeruli of STZ-induced diabetic rats from 3 days to 12 weeks after the induction of diabetes, together with histological examination. To investigate the role of prostanoids in this process, aspirin was administered in one group of diabetic rats. Furthermore, CTGF expression was analyzed in rat mesangial cells cultured under high-glucose conditions. RESULTS: Glomerular expression of CTGF and TGF-beta1 mRNA was coordinately upregulated as early as day 3, followed by fibronectin induction and mesangial matrix accumulation. Chronic aspirin treatment in diabetic rats significantly attenuated mesangial expansion, and effectively suppressed CTGF induction, as well as inhibiting the upregulation of TGF-beta1 and fibronectin expression. In cultured mesangial cells, aspirin treatment abolished high glucose-stimulated CTGF upregulation. CONCLUSIONS: These results indicate that CTGF expressed in the glomeruli is upregulated in the early stage of STZ-induced diabetic nephropathy in rats, and could be a critical mediator of the development of diabetic glomerulosclerosis. In addition, the modulatory effects of aspirin during this process suggest a role of the cyclooxygenase pathway in the progression of diabetic nephropathy.

Angiogenic protein Cyr61 is expressed by podocytes in anti-Thy-1 glomerulonephritis.

Dynamic recovery of glomerular structure occurs after severe glomerular damage in anti-Thy-1 glomerulonephritis (Thy-1 GN), but its mechanism remains to be investigated. To identify candidate genes possibly involved in glomerular reconstruction, screening was performed for genes that are specifically expressed by podocytes and are upregulated in glomeruli of Thy-1 GN. Among them, cysteine-rich protein 61 (Cyr61 or CCN1), a soluble angiogenic protein belonging to the CCN family, was identified. By Northern blot analysis, Cyr61 mRNA was markedly upregulated in glomeruli of Thy-1 GN from day 3 through day 7, when mesangial cell migration was most prominent. By in situ hybridization and immunohistochemistry, Cyr61 mRNA and protein were expressed by proximal straight tubules and afferent and efferent arterioles in normal rat kidneys and were intensely upregulated at podocytes in Thy-1 GN. Platelet-derived growth factor-BB (PDGF-BB) and transforming growth factor-beta1 (TGF-beta1), of which the gene expression in the glomeruli of Thy-1 GN was upregulated in similar time course as Cyr61, induced Cyr61 mRNA expression in cultured podocytes. Furthermore, supernatant of Cyr61-overexpressing cells inhibited PDGF-induced mesangial cell migration. In conclusion, it is shown that Cyr61 is strongly upregulated at podocytes in Thy-1 GN possibly by PDGF and TGF-beta. Cyr61 may be involved in glomerular remodeling as a factor secreted from podocytes to inhibit mesangial cell migration.

Role of connective tissue growth factor in fibronectin expression and tubulointerstitial fibrosis.

Connective tissue growth factor (CTGF) is one of the candidate factors mediating downstream events of transforming growth factor-beta (TGF-beta), but its role in fibrogenic properties of TGF-beta and in tubulointerstitial fibrosis has not yet been clarified. Using unilateral ureteral obstruction (UUO) in rats, we analyzed gene expression of TGF-beta1, CTGF, and fibronectin. We further investigated the effect of blockade of endogenous CTGF on TGF-beta-induced fibronectin expression in cultured rat renal fibroblasts by antisense oligodeoxynucleotide (ODN) treatment. After UUO, CTGF mRNA expression in the obstructed kidney was significantly upregulated subsequent to TGF-beta1, followed by marked induction of fibronectin mRNA. By in situ hybridization, CTGF mRNA was detected mainly in the interstitial fibrotic areas and tubular epithelial cells as well as in parietal glomerular epithelial cells in the obstructed kidney. The interstitial cells expressing CTGF mRNA were also positive for alpha-smooth muscle actin. CTGF antisense ODN transfected into cultured renal fibroblasts significantly attenuated TGF-beta-stimulated upregulation of fibronectin mRNA and protein compared with control ODN transfection, together with inhibited synthesis of type I collagen. With the use of a reporter assay, rat fibronectin promoter activity was increased by 2.5-fold with stimulation by TGF-beta1, and this increase was abolished with antisense CTGF treatment. Thus CTGF plays a crucial role in fibronectin synthesis induced by TGF-beta, suggesting that CTGF blockade could be a possible therapeutic target against tubulointerstitial fibrosis.

Overexpression of brain natriuretic peptide in mice ameliorates immune-mediated renal injury.

One of major causes of end-stage renal disease is glomerulonephritis, the treatment of which remains difficult clinically. It has already been shown that transgenic mice that overexpress brain natriuretic peptide (BNP), with a potent vasorelaxing and natriuretic property, have ameliorated glomerular injury after subtotal nephrectomy. However, the role of natriuretic peptides in immune-mediated renal injury still remains unknown. Therefore, the effects of chronic excess of BNP on anti-glomerular basement membrane nephritis induced in BNP-transgenic mice (BNP-Tg) were investigated and the mechanisms how natriuretic peptides act on mesangial cells in vitro were explored. After induction of nephritis, severe albuminuria (approximately 21-fold above baseline), tissue damage, including mesangial expansion and cell proliferation, and functional deterioration developed in nontransgenic littermates. In contrast, BNP-Tg exhibited much milder albuminuria (approximately fourfold above baseline), observed only at the initial phase, and with markedly ameliorated histologic and functional changes. Up-regulation of transforming growth factor-beta (TGF-beta) and monocyte chemoattractant protein-1 (MCP-1), as well as increased phosphorylation of extracellular signal-regulated kinase (ERK), were also significantly inhibited in the kidney of BNP-Tg. In cultured mesangial cells, natriuretic peptides counteracted the effects of angiotensin II with regard to ERK phosphorylation and fibrotic action. Because angiotensin II has been shown to play a pivotal role in the progression of nephritis through induction of TGF-beta and MCP-1 that may be ERK-dependent, the protective effects of BNP are likely to be exerted, at least partly, by antagonizing the renin-angiotensin system locally. The present study opens a possibility of a novel therapeutic potential of natriuretic peptides for treating immune-mediated renal injury.