Role of glycosaminoglycans in the regulation of T cell proliferation induced by thymic stroma-derived T cell growth factor

The present study investigates the regulatory effects of glycosaminoglycans such as heparin and heparan sulfate on T cell proliferation induced by thymic stromal cell monolayer or its derived T cell growth factor (TCGF). A thymic stromal cell clone (MRL104.8a) supported the growth of Ag-specific, IL-2-dependent Th cell clone (9-16) in the absence of Ag and IL-2 by producing a unique TCGF designated as thymic stroma-derived T cell growth factor (TSTGF). The addition of heparin to cultures in which the growth of 9-16 Th cells was otherwise stimulated by the MRL104.8a monolayer or a semipurified sample of the TSTGF resulted in heparin dose-dependent inhibition of 9-16 Th proliferation. The dose of heparin required for inducing 50% reduction of TSTGF-induced proliferation of Th at a given cell number was found to be proportional to the magnitude of the TSTGF added to cultures, suggesting that heparin exerted its inhibitory effect by binding to the TSTGF rather than by acting on Th cells. A similar growth-inhibiting effect of heparin was observed in IL-7-dependent proliferation of pre-B cell line or Th, but not in IL-2-dependent T cell proliferation or IL-3-dependent myeloid cell proliferation. A strong affinity of TSTGF and IL-7 for heparin was confirmed by the fact that both TSTGF and IL-7 adhered to columns of heparin-agarose and were eluted by salt. When various glycosaminoglycans were tested for the heparin-like Th growth-regulatory capacity, heparan sulfate exhibited Th growth-inhibiting ability comparable to that observed for heparin. These results indicate that the activity of thymic and/or bone marrow stroma-derived lymphocyte growth factor (TSTGF/IL-7) but not of Th-producing TCGF (IL-2) is negatively regulated by heparin or heparan sulfate, which would represent major glycosaminoglycans in the extra-cellular matrix of stromal cells.     

The complete amino acid sequence of ribonuclease from the seeds of bitter gourd (Momordica charantia)

The complete amino acid sequence of ribonuclease (RNase MC) from the seeds of bitter gourd (Momordica charantia) has been determined. This has been achieved by the sequence analysis of peptides derived by enzymatic digestion with trypsin, lysylendopeptidase, and chymotrypsin, as well as by chemical cleavage with cyanogen bromide. The protein contains 191 amino acid residues and has a calculated molecular mass of 21,259 Da. Comparison of this sequence with sequences of the fungal RNases, RNase T2, and RNase Rh, revealed that there are highly conserved residues at positions 32-38 (TXHGLWP) and 81-92 (FWXHEWXKHGTC). Furthermore, the sequence of RNase MC was found to be homologous to those of Nicotiana alata S-glycoproteins involved in self-incompatibility sharing 41% identical residues.     

A case of primary hyperparathyroidism due to an oxyphil cell adenoma.

A 59-year-old woman with primary hyperparathyroidism was found to have a parathyroid adenoma behind the left clavicle. Preoperatively, it appeared as a hypoechoic mass on ultrasonography, as a hot nodule on thallium scintigraphy, and as a high signal on T2-weighted magnetic resonance imaging. Histological, immunohistochemical and ultrastructural studies of the surgically resected tumor revealed a parathyroid adenoma composed mainly of oxyphil cells with production of a parathyroid hormone. Moreover, a multilocular lesion of lymphangiectasia was contained. Hypercalcemia was alleviated postoperatively. These observations corroborated a functioning parathyroid oxyphil cell adenoma. This is the first case report of functioning oxyphil cell adenoma of the parathyroid gland with lymphangiectasia in Japan.     

Binding characteristics of naftopidil and alpha 1-adrenoceptor antagonists to prostatic alpha-adrenoceptors in benign prostatic hypertrophy.

Binding properties of naftopidil and alpha 1-adrenoceptor antagonists to alpha-adrenoceptors in prostates from benign prostatic hypertrophy (BPH) were characterized by radioreceptor assays using [3H]prazosin and [3H]rauwolscine. Specific binding of [3H]prazosin and [3H]rauwolscine in human prostatic membranes was saturable and of high affinity, and it showed a pharmacological specificity which characterized alpha 1 and alpha 2-adrenoceptors, respectively. Naftopidil and several alpha 1 antagonists competed for prostatic [3H]prazosin binding in order: R-(-)-YM-12617 greater than prazosin greater than bunazosin greater than terazosin greater than naftopidil greater than urapidil, and the inhibitory effect (Ki = 11.6 nM) of naftopidil was 10 to 45 times less potent than quinazoline derivatives such as prazosin, bunazosin and terazosin. The potencies of these antagonists in competing for [3H]prazosin binding sites in human prostates correlated well with their pharmacological potencies (pA2). Scatchard analysis indicated that the decrease of prostatic [3H]prazosin binding by naftopidil was due to a marked increase in the Kd value without a change in the Bmax value. The inhibition of prostatic [3H]prazosin binding by naftopidil was reversible. Naftopidil also inhibited prostatic [3H]rauwolscine binding (Ki = 70.0 nM). Thus, it is suggested that naftopidil antagonizes alpha 1-adrenoceptors in human prostates in a competitive and reversible manner.     

Haematological and serum biochemical values in hairless and haired descendants of Mexican hairless dogs.

Haematological and serum biochemical measurements were carried out in 1-year-old hairless and haired hybrids derived from the Mexican hairless dog (MHD). These hybrids included F1 hybrids obtained from male MHD and female Beagles, and BCF1 hybrids obtained from male hairless F1 and female Beagles. There were no significant differences between F1 and BCF1 hybrids, nor between male and female hybrids. Except for red blood cell counts, haemoglobin concentrations and packed cell volumes which were slightly higher in MHD-descendants than in Beagles, there were no differences for haematological and serum biochemical findings between hairless and haired hybrids when compared to age-matched Beagles.     

Estimation of stature in children from second metacarpal measurements.

On the basis of 552 boys and 542 girls aged 6 to 20 years, this study examines the estimation of stature from dimensions and maturity of second metacarpals by means of linear regression equations. A combination of length and width measurements provided a more accurate estimation than each measurement individually. When taken alone, length produced a more accurate estimation than width. Sex and age factors are useful for the estimation of stature, though these variables are often unknown in the isolated bone. The samples are divided into immature and mature groups (according to skeletal maturity). Regardless of sex, stature could be estimated from the metacarpal length and width with a standard error of 4.19 cm by means of a multiple linear equation in the immature group. The mature group should be considered with adults for this purpose. Thus, taking into account their skeletal maturity, living stature could be practically estimated from the second metacarpal with significant degrees of accuracy in children.     

Estimation of age at death from second metacarpals.

This study examined the estimation of age at death from the second metacarpal in 227 individuals aged 30-98 years. Variables ascertained from each bone were: cortical thickness and microdensitometric cortical bone density measured on radiographs of the bone and total osteon count and density recorded on microradiographs of the complete cross section at its midshaft. Based on the latter two variables, two age groups were formed; a middle age group representing those individuals aged 30-65 years, and an older group aged 65+. Stepwise regression analysis of the four variables produced a series of regression equations for age estimation for the middle, old and combined age groups for each sex and sexes combined. Sex-specific equations provided better results than nonspecific ones, especially in females. Total osteon density and combined cortical thickness were found to be the most useful estimators in the middle and the old age group, respectively. The standard error of estimate was 6.71 and 6.90 years in each age group for the sexes combined. In the combined age group, age could be estimated accurately from total osteon count, cortical thickness and MD cortical bone density with the standard error of estimate of 11.10 years. The relative error of estimate ranged within +/- 30% in almost all individuals aged above 60 years.     

Production of S-lactoylglutathione by organic-solvent-extracted glyoxalase I from Hansenula mrakii

Summary Glyoxalase I was extracted from Hansenula mrakii IFO 0895 by incubating the cells with buffer solution containing 50% acetone (enzyme activity 35 units/g cells) or 50% ethyl acetate (enzyme activity 28 units/g cells) at 30°C for 10 h. Glyoxalase II was also extracted from the cells, although the activity of the enzyme was lost during incubation with organic solvents, especially at higher temperature (30°C). By using the organic-solvent-extracted fraction of H. mrakii, enzymatic production of S-lactoylglutathione was studied, and approximately 82 mmol/l (30 g/l) of S-lactoylglutathione was produced from 120 mmol/l glutathione. Offprint requests to: A. Kimura     

A sensitive nonisotopic method for the determination of intracellular azidothymidine 5'-mono-, 5'-di-, and 5'-triphosphate.

In order to analyze the efficacy of azidothymidine (AZT), it is important to know intracellular concentrations of AZT metabolites. However, it has been impossible to measure intracellular AZT 5'-monophosphate (AZT-MP), AZT 5'-diphosphate (AZT-DP), and AZT 5'-triphosphate (AZT-TP) without using isotopes. In the present study, we developed a new method to measure intracellular AZT metabolites without radiolabeled compounds. The method employed was a high-performance liquid chromatography (HPLC) system programmed for column switching technique, in which two columns were used: column 1 (TSK-G2000-SW, 300 x 7.5 mm) to preseparate AZT metabolites from major cell components, and column 2 (YMC-A-312-ODS, 150 x 6 mm) to determine the metabolites. The limit of detectability of this system was 3.3 pmol/injection. When MT-4 cells were incubated with various concentrations of AZT, intracellular concentrations of AZT-MP increased in parallel with extracellular AZT. Those of AZT-DP and AZT-TP, however, reached plateaus at 5 and 2 microM of AZT, respectively. In MT-4 and Molt-4 cells incubated with 5 microM AZT, concentrations of AZT-MP increased time dependently, while the AZT-DP/AZT-MP ratios decreased with time. These data suggest that high dose of AZT may not necessarily increase intracellular concentration of AZT-TP. The concentrations of AZT metabolites in peripheral blood mononuclear cells in a patient with AIDS and an asymptomatic carrier were measured; the concentrations were comparable to those in cultured cells. Quantitative analysis of intracellular AZT metabolites without the use of isotopes will increase safety and convenience of measurement, and take an effective step in studying pharmacokinetics of AZT in clinical materials.     

Oxidative modification of glycated low density lipoprotein in the presence of iron

This is the first observation for contributing to the glycation of low density lipoprotein (LDL) to oxidative modification of its own lipids and protein. Human plasma LDL was glycated by incubation with glucose (G-LDL). Glucose incorporated into apoprotein B was approximately 10 mol/mol of apoprotein (2.8% modification of lysine residues) and 84% of G-LDL was adsorbed on phenylboronate affinity column. G-LDL incubated with Fe3+ for 4 h caused a significantly higher level of lipid peroxidation than U-LDL (untreated with glucose), and a higher molecular weight protein was observed in apoprotein B on SDS-polyacrylamide gel electrophoresis (SDS-PAGE), increasing with incubation period. Corresponding to change on SDS-PAGE, G-LDL exposed to Fe3+ moved faster than G-LDL per se or U-LDL to anode on agarose gel electrophoresis. The higher the Fe3+ concentration, the more lipid peroxidation was caused. Alpha-tocopherol or probucol suppressed the lipid peroxidation of G-LDL exposed to Fe3+.