Variation analysis of β3-adrenergic receptor and melanocortin-4 receptor genes in childhood obesity

BACKGROUND: Decreased energy expenditure and increased food intake are principal causes for obesity. In the present study, genotypes of beta(3)-adrenergic receptor (beta(3)AR) and of melanocortin-4 receptor (MC4R), both of which are believed to have a close link to the cause of obesity, were analyzed and compared with phenotypes of childhood obesity. METHODS: Thirty-five obese children with moderate to severe obesity were enrolled. Direct sequencing of the MC4R coding region and pinpoint-polymerase chain reaction were used to detect genomic variation in the beta(3)AR gene using peripheral blood-derived DNA. RESULTS: Allele frequency of Trp64Arg variation in the beta(3)AR gene in the obese subjects was 0.16, which is comparable with that in the healthy general population in eastern Asia. Comparison of phenotypical characteristics did not show a significant difference between Trp/Trp and Trp/Arg subjects. It was notable that body height SD was significantly higher in the Trp/Trp than the Trp/Arg subjects (0.93 +/- 1.0 SD vs 0.07 +/- 1.3 SD, P= 0.03). Annual weight gains were far beyond a hypothetical fat gain in an Arg64 heterozygote with decreased energy consumption, suggesting increased food intake in childhood obesity. There was, however, no variation in the MC4R gene despite thorough sequencing of the entire coding region. CONCLUSIONS: The Trp64Arg variation in the beta(3)AR gene has no relationship to the degree or the incidence of childhood obesity. The majority of childhood obesity can be characterized as tall stature, more rapid weight gain than that expected by decreased energy expenditure. Further investigation is necessary in regard to the increased food intake as a major cause of childhood obesity.     

Inhibition of Lipid Peroxidation by Sesquiterpenoid in Heterotheca inuloides

A sesquiterpenoid, 7-hydroxy-3,4-dihydrocadalin, isolated from a Mexican medicinal plant Heterotheca inuloides was evaluated as an antioxidant. This sesquiterpenoid inhibited mitochondrial and microsomal lipid peroxidation induced by Fe(III)-ADP/NADH or Fe(III)-ADP/NADPH. Furthermore, 7-hydroxy-3,4-dihydrocadalin protected red cells against oxidative haemolysis. This sesquiterpene was thus shown to be effective in protecting biological systems against oxidative stresses.     

Cyclosporine therapy affects aminotransferase activity but not hepatitis C virus RNA levels in chronic hepatitis C

Interferon (IFN) therapy is of proven efficacy in chronic hepatitis C, but it is not universally effective and is often limited by side effects. Cyclosporine A (CsA) is a potent immunosuppressant widely used in organ transplantation. We conducted a pilot study to determine whether CsA therapy could affect aminotransferase activity and hepatitis C virus RNA levels in patients with chronic hepatitis C. Cyclosporine A was administered to 10 patients (mean age of 59 years; male: female = 9:1) who did not respond to IFN therapy previously and who had elevated serum alanine aminotransferase (ALT) values for at least 6 months. All patients were positive for HCV-RNA by RT-PCR with genotype 1b. Their mean duration of hepatitis was 15 years. Oral CsA was given for 3 months in a dose that was increased at 1 month intervals from 1.5–2.0 to 2.0–3.0 and 3.0–4.0 mg/kg per day. All patients completed the treatment schedule, although two patients developed mild non-symptomatic hypertension. Serum ALT levels gradually decreased in all but one patient. The mean percentage decrease was 59.5% at the end of therapy (from 153 ± 82 to 62 ± 48 IU/L; P < 0.02). The ALT levels fell to the normal range in five patients, although once therapy was discontinued the enzyme levels tended to return to pretreatment levels. Serum aspartate aminotransferase and g-glutamyl transpeptidase levels similarly decreased. The serum HCV-RNA titre, determined by competitive RT-PCR, did not change in any patient throughout the study period. There were no appreciable alterations in other laboratory tests, such as serum creatinine levels and lymphocyte subsets, except for an increase in serum alkaline phosphatase levels. These findings suggest that CsA, even in a relatively low dose, reduces serum aminotransferase levels without serious side effects in patients with chronic-hepatitis C, although an antiviral effect was not noted.     

A protease inhibitor attenuates gastric erosions and microcirculatory disturbance in the early period after thermal injury in rats

The effects of camostat mesilate, a synthetic serine protease inhibitor on gastric microcirculation and active oxygen species generated by leucocytes from the gastric and jugular veins in the early period after thermal injury were assessed. Male Wistar rats were anaesthetized and a 30% full skin-thickness dorsal burn was inflicted. Camostat mesilate (100 mg/kg) was dissolved in distilled water and administered orally to rats 40 min before thermal injury (the camostat group). The control animals (the vehicle group) were administered distilled water orally. Rolling leucocytes as well as Monastral blue B deposits in venules were observed using in vivo microscopy. Active oxygen species were measured by chemiluminescence. Camostat mesilate decreased the total length of gastric erosion, venular deposits of Monastral blue B, and rolling of leucocytes in venules, and relatively increased luminol-dependent chemiluminescence activity generated by zymosan-stimulated leucocytes 15 min after thermal injury. These results suggest that serine proteases are involved in the formation of gastric erosions and gastric microcirculatory disturbance in the early period after thermal injury.     

Serum levels of IL-10, IL-15 and soluble tumour necrosis factor-alpha (TNF-α) receptors in type C chronic liver disease

We previously reported that the number of TNF-α-producing cells was increased in the liver of patients with type C chronic liver disease. To understand further the pathophysiology of this change, we examined serum levels of two soluble TNF receptors, TNF-αRI (p55) and -αRII (p75), and IL-10, all of which act as TNF-α buffer, and IL-15, a novel cytokine sharing many immunological activities with IL-2, using ELISA methods. We studied control individuals and patients with type C chronic liver disease, including asymptomatic hepatitis C virus (HCV) carriers with persistently normal serum ALT values, and those with chronic hepatitis (CH), liver cirrhosis (LC) and hepatocellular carcinoma (HCC). Both types of sTNF-αR closely correlated with disease progression. Patients with LC and HCC had significantly elevated levels for sTNF-αRII compared with the other patient groups and controls. Serum IL-10 levels were significantly greater in all chronic liver disease groups than in controls. With respect to IL-15, the values were high in CH, LC and HCC compared with those of controls. Notably, HCC patients showed highest values for both IL-10 and IL-15, with significant differences from the other patient groups. Serial determinations revealed that interferon (IFN) treatment for CH patients resulted in the suppression of circulating IL-10 and IL-15 levels along with decrease in serum aminotransferase values. Both cytokines remained at decreased levels after cessation of therapy in patients who went into clinical and virological remission. On the other hand, treatment did not affect serum levels of sTNF-αRs. These findings indicate that serum levels of these molecules correlated with disease progress in chronic HCV infection, and that IL-10 and IL-15 may reflect the degree of inflammation in the liver. It is also suggested that both cytokines may be related to the development of HCC.     

Modulation of Fas-mediated apoptosis of human thyroid epithelial cells by IgG from patients with Graves' disease (GD) and idiopathic myxoedema

The expression of two autoimmune thyroid diseases, GD and idiopathic myxoedema, is associated with antibodies to the thyroid-stimulating hormone (TSH) receptor. Thyroid stimulating antibodies (TSAb) in GD are TSH agonists and cause hyperthyroidism as well as goitre, whereas thyroid stimulation blocking antibodies (TSBAb) in idiopathic myxoedema are TSH antagonists and cause hypothyroidism and thyroid atrophy. We investigated the effect of antibodies to TSH receptor on Fas-mediated apoptosis of thyroid epithelial cells (thyrocytes). Human IgG was isolated from healthy donors, patients with GD and idiopathic myxoedema. Human thyrocytes were obtained from surgical specimens. Thyrocytes were cultured in the presence or absence of human IgG with or without interferon-gamma (IFN-γ) or IL-1β for a specified time. After incubation, we examined the level of cAMP in cultured supernatants and both Fas and Bcl-2 expression on thyrocytes. In addition, we examined anti-Fas-mediated apoptosis of thyrocytes. Fas expression on thyrocytes was significantly down-regulated by Graves' IgG and TSH, although idiopathic myxoedema IgG did not affect Fas expression on thyrocytes. Idiopathic myxoedema IgG abrogated the effect of TSH on both cAMP production and inhibition of Fas expression on thyrocytes. Treatment of thyrocytes with IL-1β or IFN-γ caused a marked augmentation of Fas expression on thyrocytes. The increase of Fas expression of thyrocytes induced by IL-1β or IFN-γ was significantly suppressed in the presence of TSH or Graves' IgG. Anti-Fas-induced apoptosis of thyrocytes was observed in thyrocytes treated with IL-1β or IFN-γ, but was markedly inhibited in the presence of TSH or Graves' IgG. Furthermore, idiopathic myxoedema IgG abrogated most of the inhibitory effect of TSH on Fas-mediated apoptosis of thyrocytes treated with IL-1β or IFN-γ. Bcl-2 expression of thyrocytes did not change after stimulation with TSH, Graves' IgG, idiopathic myxoedema IgG, IL-1β or IFN-γ. These results suggest that TSAb found in Graves' patients may be potentially involved in the development of goitre by inhibition of Fas-mediated apoptosis of thyrocytes. In addition, TSBAb inhibit the action of TSH and increase the sensitivity toward Fas-mediated apoptosis of thyrocytes, inducing thyroid atrophy seen in patients with idiopathic myxoedema.     

Atypical polypoid adenomyomas of the uterus

We performed a clinicopathological immunohistochemical, ultrastructural, and flow cytometric study on six cases of atypical polypoid adenomyoma of the uterus including one with an adenocarcinoma within it. The tumours occurred in nulliparous women aged 22–48 years (average, 33.0 years); three arose in the uterine corpus, and three in the endocervix. Histologically, they were composed of endometrial glands admixed with a stromal component of interlacing bundles of smooth muscle cells. The glands exhibited varying degrees of architectural and cytological atypia. Most of the stromal cells showed strong staining for HHF35, alpha-smooth muscle actin, and vimentin, and some cells contained desmin. Electronmicroscopy, in one case, confirmed the presence of a well-differentiated smooth muscle component. The stromal component may arise as a result of extensive metaplasia of endometrial stromal cells. Uninvolved endometrium showed ciliated cell metaplasia in three patients, and atypical complex hyperplasia in two. One patient had a well-differentiated adenocarcinoma of endometrioid type arising in an endocervical atypical polypoid adenomyoma. All tumours had a diploid DNA content and relatively small S phase fraction (average, 6.23%). The follow-up periods ranged from 4 to 42 months (average, 13.5 months), and all patients were alive and well. Although the histogenesis of atypical polypoid adenomyoma of the uterus remains uncertain, it is suggested that it may arise because of oestrogen-related factors.     

Importance of Hydrostatic Pressure and Irrigation for Hemostasis in Neuroendoscopic Surgery

Recently neurosurgical operations have been carried out with water irrigation such as endoscopic third ventriculostomy and tumor resections in ventricles. Water irrigation is one of several published methods that promote hemostasis; however, not enough experimental evidence exists on its efficacy. In this study, we investigate whether hydrostatic pressure and persistent irrigation promote hemostasis in neuroendoscopic surgery. We dissected tails of 12–16-week-old C57BL/6 male mice at 5 mm proximal from the tip and checked for bleeding times under dry and wet conditions at pressures of 0 cmH₂O, 10 cmH₂O, 15 H₂O, and 20 cmH₂O without persistent irrigation to bleeding point and 10 cmH₂O with persistent irrigation. We then examined the dissected edge with hematoxylin–eosin staining and measured the size of vessels. The average bleeding time of each group is as follows: dry: 203.4 sec, wet: 164.4 sec, 5 cmH₂O: 138.6 sec, 10 cmH₂O: 104.6 sec (P <0.001), 20 cmH₂O: 56 sec (P <0.001), and 10 cmH₂O with persistent irrigation: 72.8 sec (P <0.01 compared to 10 cmH₂O without persistent irrigation). The maximum caliber of mice’s tail artery was 50–60 μm. Hydrostatic pressure and irrigation are important factors contributing to hemostasis.     

Vitamin D-dependent rickets type I and type II

Two distinct hereditary defects, vitamin D-dependent rickets type I (VDDR I) and type II (VDDR II), have been recognized in vitamin D metabolism. VDDR I is suggested to be a deficiency of the renal 25-hydroxyvitamin D (25(OH)D)-1α-hydroxylase. Muscle weakness and rickets are the prominent clinical findings. A normal physiologic dose of 1α-hydroxyvitamin D₃ and 1,25-dihydroxyvitamin D₃ is sufficient to maintain remission of rickets in this disorder. VDDR II consists of a spectrum of intracellular vitamin D receptor (VDR) defects and is characterized by the early onset of severe rickets and associated alopecia. This can be attributed to mutations in the VDR gene. Massive doses of vitamin D analogs and calcium supplementation is usually required for the treatment; however, the response to therapy is sometimes variable.     

Keratinocyte growth factor is an endogenous stimulant of rabbit gastric epithelial cell proliferation and migration in primary culture

Mesenchymal-epithelial interactions are important in the gastric mucosal repair. However, specific factors responsible for such interactions have not been established. In the present study, keratinocyte growth factor (KGF) significantly stimulated proliferation of gastric epithelial cells dose dependently and synergistically with hepatocyte growth factor (HGF), epidermal growth factor (EGF) and insulin. Restitution of gastric epithelial monolayers was also assessed, using a round wound restitution model. Keratinocyte growth factor facilitated the restitution of gastric epithelial cells significantly but did not have any effects on gastric fibroblasts. Keratinocyte growth factor receptor mRNA was expressed by gastric epithelial cells, indicating that these effects were elicited by the specific receptor mediated pathway. Northern blot analysis revealed the expression of KGF mRNA in gastric fibroblasts but not in gastric epithelial cells, indicating the production of KGF. These results suggest that KGF might be involved in gastric mucosal repair, through mesenchymal-epithelial interaction.