Hydroxychloroquine modulates elevated expression of S100 proteins in systemic lupus erythematosus.

OBJECTIVES: We investigated the effect of hydroxychloroquine (HCQ) on S100A8 and S100A9 serum levels in systemic lupus erythematosus (SLE) patients with low disease activity receiving immunosuppressants. METHODS: SELENA-SLEDAI, Cutaneous Lupus Erythematous Disease Area and Severity Index (CLASI) and serum levels of complement factors, anti-dsDNA antibodies, and white blood cell, lymphocyte, and platelet counts were used to evaluate disease activity, cutaneous disease activity, and immunological activity, respectively. Serum S100A8 and S100A9 were measured at HCQ administration and after 3 or 6 months using ELISA. RESULTS: S100A8 and S100A9 serum levels were elevated at baseline and the magnitude of decrease from baseline at 3 and 6 months after HCQ administration was greater in patients with renal involvement than in those without (baseline: S100A8, p = 0.034; S100A9, p = 0.0084; decrease: S100A8, p = 0.049; S100A9, p = 0.023). S100 modulation was observed in patients with (n = 17; S100A8, p = 0.0011; S100A9, p = 0.0002) and without renal involvement (n = 20; S100A8, p = 0.0056; S100A9, p = 0.0012), and was more apparent in patients with improved CLASI activity scores (improved: S100A8, p = 0.013; S100A9, p = 0.0032; unimproved: S100A8, p = 0.055; S100A9, p = 0.055). No associations were observed for immunological biomarkers. CONCLUSION: HCQ may improve organ involvement in SLE by modulating S100 protein levels, especially in patients with renal or skin involvement.

Facet cyst haematoma in the lumbar spine: a report of four cases.

We present 4 cases of facet cyst haematoma in the ligamentum flavum of the lumbar spine. All patients presented with a one-to-3-month history of back pain or numbness in the legs, and sudden neurological deterioration. One also developed cauda equina syndrome and another developed radiculopathy. In all cases, magnetic resonance imaging showed a mass with high signal intensity on both T1- and T2-weighted images. Facet arthrography and computed tomography revealed communication between the mass and the neighbouring facet joint. The haematomas were removed en bloc with the ligamentum flavum. They were surrounded by the ligament and contained degenerated and lacerated elastic fibres but no synovial lining cells. Facet cyst haematoma is so-named because of bleeding from tissue adjacent to the facet joint into a pre-existing facet cyst.

Increased expression of intercellular adhesion molecule-1 (ICAM-1) in diabetic rat glomeruli: glomerular hyperfiltration is a potential mechanism of ICAM-1 upregulation.

Mononuclear cells, including monocytes/macrophages and T-cells, are considered to be involved in the progression of diabetic nephropathy, although the mechanism of their recruitment into diabetic glomeruli is unclear. The intercellular adhesion molecule-1 (ICAM-1) promotes the infiltration of leukocytes into atherosclerotic lesions as well as inflammatory tissues. In the present study, we investigated the expression of ICAM-1 in the glomeruli of streptozotocin-induced diabetic rats. The expression of ICAM-1 was increased significantly during the early stage of diabetes. The number of mononuclear cells, primarily monocytes/macrophages and lymphocytes, was significantly increased in diabetic glomeruli. Mononuclear cell infiltration into diabetic glomeruli was prevented by anti-ICAM-1 monoclonal antibody. Insulin treatment decreased ICAM-1 expression and mononuclear cell infiltration. The ICAM-1 expression on cultured human umbilical vein endothelial cells was not induced under high glucose culture conditions. Glomerular hyperfiltration is a characteristic change in the early stage of diabetic nephropathy. Treatment with aldose reductase inhibitor, which prevented glomerular hyperfiltration without changes in blood glucose levels, decreased ICAM-1 expression and mononuclear cell infiltration. Moreover, we examined the ICAM-1 expression in the glomeruli of the 5/6 nephrectomized rat, which is a model for glomerular hyperfiltration without hyperglycemia. The ICAM-1 expression and infiltration of mononuclear cells was significantly increased in the glomeruli of 5/6 nephrectomized rats. We conclude that ICAM-1 is upregulated and promotes the recruitment of mononuclear cells in diabetic glomeruli. Moreover, glomerular hyperfiltration that occurs in the early stage of diabetic glomeruli may be one of the potential mechanisms of ICAM-1 upregulation in diabetic nephropathy.

Ultrastructure of mesangial type III collagen deposition in a patient with IgA nephropathy.

We report a case of IgA nephropathy with focal and segmental deposition of type III collagen in mesangium, confirmed by immunohistochemical and electron microscopic methods. Tissue negative staining showed that focal and segmental fibrotic lesions in the mesangial area consisted of disarrayed or curled striated collagen fibers and striated membranous structures. Diabetes mellitus, hypertension, and advanced glomerular sclerosis were absent in this case, and mesangial cells surrounding the type III collagen showed vacuolar degeneration revealed by electron microscopy. Production of type III collagen may be the marker for phenotypic change of mesangial cells in immune-mediated glomerular diseases.

Specific chemical cleavage of asparaginyl and glycyl-glycine bonds in peptides and proteins by anhydrous hydrazine vapor.

Hydrazinolysis of peptide or protein has been used for C-terminal amino acid determination by Akabori et al. (1952). In this study, proteins were reacted with anhydrous hydrazine vapor at 20 degrees C for 16 h. Asparaginyl linkages were cleaved. Asparagine and glutamine were converted to their hydrazides, beta-hydrazidyl aspartic acid and gamma-hydrazidyl glutamic acid, respectively, even under milder conditions. The former hydrazide cyclizes to a 6-membered ring, asparaginyl bond at the carboxyl side. Other cleavages, including the glycyl-glycine bond, were also observed.

Steroid hormones differentially induce transcription of the chicken ovalbumin gene, but stabilize the mRNA with the same half-life.

The stabilization of chicken ovalbumin (OVA) mRNA by different classes of steroid hormones (estrogen, progesterone, glucocorticoid, and androgen) was studied in the oviducts of chicks treated with combinations of four steroids. The combination of estrogen with progesterone, glucocorticoid, or androgen enhanced the induction of the OVA gene more than did estrogen alone. Run-on analysis of the isolated oviduct nuclei to measure the transcription rate of the OVA gene showed that the enhanced induction of the OVA gene by the combined hormone treatments was partly caused by an increased rate of transcription. The half-life of OVA mRNA as determined using a transcription inhibitor (actinomycin D) was estimated to be about 24 h irrespective of the hormone treatment, though the half-life was about 6 h in the absence of hormones. These results suggested that the prolongation of the half-life of OVA mRNA by steroid hormones is constant irrespective of differential transcription rates of the OVA gene.

Resolution of Fusarium sporotrichioides Proteins by Two-Dimensional Polyacrylamide Gel Electrophoresis and Identification by Sequence Homology Comparison in Protein Data Base.

Proteins from Fusarium sporotrichioides M-1-1, a T2-toxin-producing strain, were separated by two-dimensional polyacrylamide gel electrophoresis. One thousand two hundred and forty-four protein spots were resolved and 103 protein spots were subjected to N-terminal sequencing. Fifty-eight protein spots were sequenced and 48 proteins were observed to have blocked N termini. Forty out of 58 sequenced proteins were identified by homology search against the PIR protein sequence data base and protein superfamily data base, while the residual 18 sequences were not identified. Twenty-seven of the N-terminal-blocked proteins were subjected to mild anhydrous hydrazine vapor deblocking. Twenty-four spots were not deblocked indicating the presence of acyl groups at the N termini, while 3 proteins were deblocked showing the blocked group to be pyrroglutamyl carboxylic acid residues. The results can provide a more global view of cellular genetic expression than any other technique. The created data may offer a unique opportunity to link information with DNA sequence data. Copyright 1995 S. Karger AG, Basel

Long-term administration of adrenocorticotropin modulates the expression of IGF-I and TGF-beta 1 mRNAs in the rat adrenal cortex.

The effects of long-term adrenocorticotropin (ACTH) therapy on the expression of IGF-I and TGF-beta 1 on rat adrenal cortex was investigated. ACTH (0.1 mg/kg/day) or saline as control was injected intraperitoneally in 5-week-old Wistar rats every day for 4 weeks. ACTH significantly increased adrenal weight (P < 0.05) and serum corticosterone (P < 0.05). Competitive RT-PCR analysis on the adrenocortical mRNA showed increased IGF-I (P < 0.01) at 4 weeks of ACTH and increased TGF-beta 1 (P < 0.01) at 1 week of ACTH compared the control group. ACTH also significantly increased proliferating cell nuclear antigen mRNA level (P < 0.01), at 4 weeks of treatment, which correlated with IGF-I level (P < 0.01), but correlated negatively with ACTH-stimulated TGF-beta 1 level (P < 0.05). There was a weak correlation between IGF-I and serum corticosterone (P < 0.05), and between TGF-beta 1 mRNA levels and serum corticosterone concentration (P < 0.05). Histologically, ACTH induced hypertrophy in the zona fasciculata cells and increased the clear cells containing lipid deposits. Immunohistochemistry showed that IGF-I peptide was mainly expressed in the periphery of the zona fasciculata at 4 weeks of ACTH therapy, while the same therapy caused a slight increase in TGF-beta 1 expression in the same area. Our results show that an increase in adrenocortical growth resulting from ACTH treatment is associated with an increase in IGF-I mRNA expression but only a transient increase in TGF-beta 1 mRNA expression.

Clinical evaluation of muscle strength in 20-79-years-old obese Japanese.

It is well known that obesity is closely related to non-insulin-dependent diabetes mellitus, hyperlipidemia, hypertension and cardiovascular disease, and the insulin resistance associated with obesity is supposed to play a central role for the development of these diseases. Thus, effective prevention and treatment of obesity need to be explored. In 357 obese (body mass index > or =26.4) subjects, aged 20-79 years, grip and leg strength were determined and compared with age- and sex-matched 1683 nonobese control subjects. Age-dependent alteration of body composition, evaluated by waist-hip ratio and the relative fat mass volume, was also compared. Finally, the relationship between the number of risk factors related to atherosclerosis and muscle strength was evaluated. Grip and leg strength in obese subjects were obviously stronger than controls under the age of 60 in both sexes. However, in the subjects over 60 years old, muscle strength was similar between obese subjects and controls. Weight bearing index (WBI) (leg strength (kg)/body weight (kg)) in obese subjects was remarkably lower than that in controls in all generations. In obese subjects, the waist-hip ratio and relative percentage of fat increased with aging, and obese subjects with multiple risk factors had higher waist-hip ratio and a tendency for lower muscle strength. Reduced WBI was considered to be a fundamental feature of obese subjects, and obese subjects increased fat composition with aging, which may be linked with low muscle strength. Thus, we need to design the most effective protocols to maximize and maintain quantitative and qualitative properties of muscle.

Nitric oxide system is involved in glomerular hyperfiltration in Japanese normo- and micro-albuminuric patients with type 2 diabetes.

Glomerular hyperfiltration plays a pathogenic role in the early stages of diabetic nephropathy. Experimental studies in laboratory animals suggest that nitric oxide (NO) might be involved in the pathogenesis of glomerular hyperfiltration. We performed a cross-sectional study to determine the relationship between diabetic glomerular hyperfiltration and the NO system. Normoalbuminuric (n=41), microalbuminuric (n=25), and macroalbuminuric (n=16) patients with type 2 diabetes were recruited in this study and compared with age-matched 84 non-diabetic control subjects. Creatinine clearance and urinary NO(2)(-)/NO(3)(-) excretion (urinary NOx) were measured, and the expression of endothelial cell nitric oxide synthase (ecNOS) was evaluated in human renal tissues. Glomerular hyperfiltration was present in 19 (37.5%) and nine (36.6%) of normoalbuminuric and microalbuminuric type 2 diabetic patients, respectively. The urinary NOx was significantly higher in normoalbuminuric patients compared with normal subjects. Creatinine clearance correlated significantly with urinary NOx in normoalbuminuric and microalbuminuric patients. Immunohistochemical staining intensities for ecNOS were significantly increased in glomerular endothelial cells of microalbuminuric type 2 diabetic patients as compared with the control subjects. These results suggest that NO may contribute to the pathogenesis of glomerular hyperfiltration in Japanese type 2 diabetic patients.

Re-evaluation of exercise prescription for Japanese type 2 diabetic patients by ventilatory threshold.

Prescription of aerobic exercise for Type 2 diabetes mellitus (Type 2 DM) in clinical practice is frequently based on exercise intensity at maximum heart rate (60

Evaluation of visceral adipose accumulation in Japanese women and establishment of a predictive formula.

Abdominal obesity is a known risk factor for diabetes-related diseases. This study aimed to establish a formula to predict visceral abdominal fat area on the basis of simple clinical and anthropomorphic parameters easily measured in the clinic. We determined visceral fat (V) and subcutaneous fat (S) areas in 115 Japanese women using the standard procedure based on computed tomography (CT) at umbilical level. Furthermore, we measured clinical and anthropometric parameters including height, weight, waist circumference, hip circumference, skin fold thickness and body fat percentage. In 115 subjects, V area was 87.8+/-52.5 cm2 and S area was 221.1+/-99.7cm2. Abdominal obesity is diagnosed in Japan as a V area > or =100 cm2; on this basis 42 women (37%) had abdominal obesity. The prevalences of diabetes and related diseases were significantly higher among women with abdominal obesity. By simple regression analysis, V and S areas significantly correlated with anthropometric parameters: in particular, V area correlated with waist circumference (r=0.745, p<0.01) and S area with body mass index (r=0.793, p<0.01). However, these parameters were not sufficient to predict V area. By multiple regression analysis using simple parameters, we established the following formula to predict visceal fat: V area = 159.475 + 1.023(age) - 2.119(height) + 1.454(body weight) + 2.841(waist circumference) - 1.208(hip circumference) (r=0.812, p<0.01). The V area calculated by formula correlated (r=0.761) with that determined by CT in a second age-matched group of 31 Japanese women. The present study confirms that visceral adipose tissue is closely associated with type 2 diabetes mellitus, dyslipidemia and hypertension, and generated a formula to predict visceral adipose tissue accumulation.

Clinical evaluation of physical fitness in male obese Japanese.

OBJECTIVE: To establish an effective exercise prescription, the exercise capacity of obese subjects was investigated. METHODS: In 71 male obese (body mass index: BMI > or = 26.4) Japanese, aged 26-77 years, the body composition and physical fitness such as aerobic exercise level, muscle strength and flexibility were determined and compared with those of age- and sex-matched 71 borderline (24.2 < or = BMI < 26.4) and 71 normal Japanese men (19.8 < or = BMI < 24.2). The relationship between body composition and other parameters was evaluated in obese subjects. RESULTS: In obese subjects, body weight, BMI, body fat percentage, waist hip ratio, and skinfold thickness were higher than those of borderline and normal subjects. Lower aerobic exercise level and weight bearing index (WBI) were noted in obese subjects. There was a negative correlation between waist hip ratio and aerobic exercise level (oxygen uptake at ventilatory threshold) (r = -0.509, P < 0.01) and also between waist hip ratio and WBI (r = -0.315, P < 0.01). CONCLUSIONS: Lower physical fitness is considered to be a general characteristic of male obese Japanese, and the accumulation of visceral adipose tissue might affect exercise capacity.

[Stability of phenyl salicylate in alcoholic-aqueous solution].

The stability and degradation pathways of phenyl salicylate (PS) in 50 v/v% aqueous ethanol were investigated over the pH range of 3.2-8.4. The experiments were performed at three different temperatures (5, 23 and 50 degrees C) for four months by the use of high performance liquid chromatography. The degradation of PS was observed as a pseudo first-order kinetics not only in the alkaline region but in the acidic region and shown to proceed via transesterification (k1) and hydrolysis (k2) processes. The transesteified product, ethyl salicylate (ES), further underwent hydrolysis (k3) (k1 > k2 > k3). On the basis of kinetic studies, it was found that the degradation rate of PS decreased with decreasing pH and temperature. The half-lives of PS at pH 6.3 were 6.6 d (50 degrees C), 178 d (23 degrees C) and 2934 d (5 degrees C), respectively, and PS was stable at below ca. pH 4. On the other hand, PS in 100% ethanol also underwent the breakdown to ES by k1 process and its half-lives were 2.5 d (50 degrees C), 31.5 d (23 degrees C) and 257 d (5 degrees C), respectively. The present results is useful for the integrity of pharmaceutical formulations and delivery profiles.

[Determination of sennoside A and sennoside B in formulation using capillary electrophoresis].

The determination of sennoside A (SA) and sennoside B (SB) by capillary zone electrophoresis was developed. The separation of SA and SB was performed in 100 mM of the 3-[cyclohexylamino]-1-propanesulfonic acid (CAPS) buffer (pH 10.0), and the migration time of SA and SB was found to be both less than 7 min. This method was applied to the analyses of seven commercial formulations containing SA and SB without previous treatment. The statistical comparison of the results obtained from both capillary electrophoresis and HPLC methods revealed an absolute correlation.

Comparison of air displacement plethysmograph and bioelectrical impedance for assessing body composition changes during weight loss in Japanese women.

OBJECTIVE: To investigate the difference between air displacement plethysmograph (ADP) and bioelectrical impedance (BI) for assessing body composition changes during weight loss in Japanese women. DESIGN: Cross-sectional and longitudinal investigation study. SUBJECTS: In a first cross-sectional study, we used data of 7380 Japanese women aged 14-83 years [body mass index (BMI): 22.6 +/- 3.7 kg/m(2)] at baseline. Among the participants, we used data of 54 women (BMI: 25.9 +/- 3.9 kg/m(2)) for a 5-month follow-up study. MEASUREMENTS: Body fat percentage was evaluated by ADP and BI, and body fat (kg) and lean body mass (kg) were calculated. Anthropometric parameters, such as height, weight, BMI, waist circumference, hip circumference and waist hip ratio, were also measured. In a second longitudinal study, they were also investigated before and after the study. RESULTS: In a first cross-sectional study, body fat percentage was 31.2 +/- 7.0% in ADP and 25.7 +/- 6.8% in BI. A significant relationship was observed between ADP and BI (r = 0.761, p < 0.01). In a second longitudinal study, body weight was significantly reduced. There were significant method (ADP vs. BI) and time (pre and post) effects but no interactions in assessing body fat percentage. There was a significant relationship between ADP and BI in body fat percentage, body fat and lean body mass. Delta body fat percentage (r = 0.386, p < 0.01) and delta body fat (r = 0.556, p < 0.01) measured by ADP were weakly correlated with those measured by BI. However, delta lean body mass measured by ADP was not correlated with that measured by BI. CONCLUSION: There was a discrepancy between ADP and BI for assessing body composition changes in Japanese women. ADP is preferentially recommended for evaluating body composition.

Re-evaluation of exercise intensity for overweight Japanese men by ventilatory threshold.

AIM: To re-evaluate exercise intensity for overweight Japanese men by ventilatory threshold (VT). METHODS: Cross sectional clinical intervention study. SUBJECTS AND MATERIALS: One hundred and ten overweight Japanese men aged 32-59 years were recruited. The average body mass index was 28.5+/-2.5 kg/m(2). Aerobic exercise level was evaluated by measuring VT. Fifty per cent heart rate (HR) reserve by Karvonen's formula was calculated and compared with HR at VT. Fat distribution was evaluated by visceral fat and subcutaneous fat areas measured with computed tomography scanning at umbilical levels. Anthropometric parameters such as height, body weight and body fat percentage were also measured. RESULTS: There was significant correlation between HR at VT and 50% HR reserve by Karvonen's formula (r=0.642, p<0.01). However, HR at VT was significantly lower than 50% HR reserve by Karvonen's formula (HR at VT 103.6+/-10.6 beat/min vs. 50% HR reserve 125.0+/-7.1 beat/min, p<0.01) and HR at VT was corresponded to 28.5+/-8.3% HR reserve by Karovonen's formula. CONCLUSION: The present study indicated that exercise intensity of 30% HR reserve should be recommended for overweight Japanese men.

A new air displacement plethysmograph for the determination of Japanese body composition.

A new device based on the plethysmographic measurement of body volume, named the BOD POD Body Composition System, was developed for the purpose of estimating body composition. The performance, reliability, validity, and clinical application of this system were evaluated in Japanese subjects. The coefficient variation (CV:%) in same-day tests was 2.48, in three separate-day tests it was 2.27, and for independent operators it was 4.53, respectively. There was a clear correlation between the results from BOD POD and those from dual energy X-ray absorptiometry (DEXA) (r = 0.910, p < 0.01). Finally, the fat body composition of 4489 subjects (1499 men, 2990 women) was clinically measured by BOD POD. The findings indicate that the BOD POD is a highly reliable and valid method for determining body fat percentage. This new method has several advantages, e.g. it is quick, simple to operate and may accommodate wide populations.

Separation and characterization of Arabidopsis thaliana proteins by two-dimensional gel electrophoresis.

Arabidopsis (Arabidopsis thaliana) proteins were isolated from five tissues (leaf, stem, root, seed and callus), and separated by two-dimensional gel electrophoresis (2-DE). 2-DE was carried out by immobilized pH gradient (IPG) in the first dimension, and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in the second dimension. With the aid of comigrated five-marker proteins, the patterns of 2-DE gels for each tissue were graphically combined by a computer into a single synthetic image for the integrated Arabidopsis protein spots. The protein spot images, altogether 4763, were characterized by both molecular mass and isoelectric point. Partial amino(N)-terminal sequences of 101 protein spots were analyzed by Edman degradation. Fifty seven proteins were partially sequenced and 46 proteins appeared to have blocked N-termini. Deblocking by hydrazine vapor was carried out on 14 proteins and two of them were found to be pyroglutamyl-blocked N-termini. Forty seven new proteins were found by the present investigation.