Flow cytometric analysis of cell cycle kinetics of anagen scalp hair correlated with plasma androgens in hirsutism

Cell kinetics of anagen scalp hair bulbs obtained from hirsute (n = 13) as well as healthy (n = 10) females were analysed by DNA-flow cytometry. The cell cycle kinetics in hirsutism revealed a significant increase of S-phase cells (10.2%) and a significant decrease of G0/1-phase cells (80.7%) compared with healthy females (S-phase 7.5%, G0/1 phase 86%). Moreover, dehydroepiandrosterone sulfate (DHEA-S) levels and cell cycle kinetics obtained from the hirsute females yielded a strong correlation between the height of S-phase percentages and DHEA-S values, whereas no correlation could be proved between testosterone levels and DNA-FCM data. Therefore, the weak androgen DHEA-S is assumed to be one hormonal factor influencing the cellular growth kinetics of hair bulbs in androgen-sensitive scalp areas.     

Synthesis of phosphatidylglycerol by chloroplasts from leaves of Spinacia oleracea L. (spinach)

Purified chloroplasts from leaves of Spinacia oleracea L. (spinach) incorporated glycerol 3-phosphate into diacylglycerol, monoacylglycerol, phosphatidylglycerol, phosphatidic acid, and lysophosphatidic acid. The omission of ATP or CTP, CoA or illumination decreased the incorporation markedly. The fraction of incorporated glycerol 3-phosphate found in phosphatidylglycerol was greatly reduced by the omission of bicarbonate, acetate, and ATP, or in darkness, low-osmolarity medium, or high magnesium ion concentration (10 mM). Incorporation of glycerol 3-phosphate into lipid and specifically into phosphatidylglycerol was optimal at a $\text{Mg}{}^{\mathrm{2+}}\text{CTP}$ ratio of 1, whereas the optimal ratio for $\text{Mg}{}^{\mathrm{2+}}\text{ATP}$ was closer to 2. The $\text{Mg}{}^{\mathrm{2+}}\text{CTP}$ gave lower total incorporation but a higher fraction of incorporation in phosphatidylglycerol. Triton X-100 inhibited incorporation of glycerol 3-phosphate into lipid, especially into phosphatidylglycerol.     

Angelman syndrome and severe infections in a patient with de novo 15q11.2–q13.1 deletion and maternally inherited 2q21.3 microdeletion

Angelman syndrome is a neurodevelopmental disorder characterized by mental retardation, severe speech disorder, facial dysmorphism, secondary microcephaly, ataxia, seizures, and abnormal behaviors such as easily provoked laughter. It is most frequently caused by a de novo maternal deletion of chromosome 15q11–q13 (about 70–90%), but can also be caused by paternal uniparental disomy of chromosome 15q11–q13 (3–7%), an imprinting defect (2–4%) or in mutations in the ubiquitin protein ligase E3A gene UBE3A mostly leading to frame shift mutation. In addition, for patients with overlapping clinical features (Angelman-like syndrome), mutations in methyl-CpG binding protein 2 gene MECP2 and cyclin-dependent kinase-like 5 gene CDKL5 as well as a microdeletion of 2q23.1 including the methyl-CpG binding domain protein 5 gene MBD5 have been described. Here, we describe a patient who carries a de novo 5 Mb-deletion of chromosome 15q11.2–q13.1 known to be associated with Angelman syndrome and a further, maternally inherited deletion 2q21.3 (~ 364 kb) of unknown significance. In addition to classic features of Angelman syndrome, she presented with severe infections in the first year of life, a symptom that has not been described in patients with Angelman syndrome. The 15q11.2–q13.1 deletion contains genes critical for Prader–Willi syndrome, the Angelman syndrome causing genes UBE3A and ATP10A/C, and several non-imprinted genes: GABRB3 and GABRA5 (both encoding subunits of GABA A receptor), GOLGA6L2, HERC2 and OCA2 (associated with oculocutaneous albinism II). The deletion 2q21.3 includes exons of the genes RAB3GAP1 (associated with Warburg Micro syndrome) and ZRANB3 (not disease-associated). Despite the normal phenotype of the mother, the relevance of the 2q21.3 microdeletion for the phenotype of the patient cannot be excluded, and further case reports will need to address this point.     

Kinetics of IL-7 and IL-15 Levels after Allogeneic Peripheral Blood Stem Cell Transplantation following Nonmyeloablative Conditioning

Background

We analysed kinetics of IL-7 and IL-15 levels in 70 patients given peripheral blood stem cells after nonmyeloablative conditioning.

Methods

EDTA-anticoagulated plasma and serum samples were obtained before conditioning and about once per week after transplantation until day 100. Samples were aliquoted and stored at −80°C within 3 hours after collection until measurement of cytokines. IL-7 and IL-15 levels were measured by ELISAs.

Results

Median IL-7 plasma levels remained below 6 pg/L throughout the first 100 days, although IL-7 plasma levels were significantly higher on days 7 (5.1 pg/mL, P = 0.002), 14 (5.2 pg/mL, P<0.001), and 28 (5.1 pg/mL, P = 0.03) (but not thereafter) than before transplantation (median value of 3.8 pg/mL). Median IL-15 serum levels were significantly higher on days 7 (12.5 pg/mL, P<0.001), 14 (10.5 pg/mL, P<0.001), and 28 (6.2 pg/mL, P<0.001) than before transplantation (median value of 2.4 pg/mL). Importantly, IL-7 and IL-15 levels on days 7 or 14 after transplantation did not predict grade II–IV acute GVHD.

Conclusions

These data suggest that IL-7 and IL-15 levels remain relatively low after nonmyeloablative transplantation, and that IL-7 and IL-15 levels early after nonmyeloablative transplantation do not predict for acute GVHD.     

Sodium sensing induces different changes in free cytosolic calcium concentration and pH in salt-tolerant and -sensitive rice (Oryza sativa) cultivars

Perception of salt stress in plant cells induces a change in the free cytosolic Ca²⁺, [Ca²⁺]_cyt, which transfers downstream reactions toward salt tolerance. Changes in cytosolic H⁺ concentration, [H⁺]_cyt, are closely linked to the [Ca²⁺]_cyt dynamics under various stress signals. In this study, salt‐induced changes in [Ca²⁺]_cyt, and [H⁺]_cyt and vacuolar [H⁺] concentrations were monitored in single protoplasts of rice (Oryza sativa L. indica cvs. Pokkali and BRRI Dhan29) by fluorescence microscopy. Changes in cytosolic [Ca²⁺] and [H⁺] were detected by use of the fluorescent dyes acetoxy methyl ester of calcium‐binding benzofuran and acetoxy methyl ester of 2′, 7′‐bis‐(2‐carboxyethyl)‐5‐(and‐6) carboxyfluorescein, respectively, and for vacuolar pH, fluorescent 6‐carboxyfluorescein and confocal microscopy were used. Addition of NaCl induced a higher increase in [Ca²⁺]_cyt in the salt‐tolerant cv. Pokkali than in the salt‐sensitive cv. BRRI Dhan29. From inhibitor studies, we conclude that the internal stores appear to be the major source for [Ca²⁺]_cyt increase in Pokkali, although the apoplast is more important in BRRI Dhan29. The [Ca²⁺]_cyt measurements in rice also suggest that Na⁺ should be sensed inside the cytosol, before any increase in [Ca²⁺]_cyt occurs. Moreover, our results with individual mesophyll protoplasts suggest that ionic stress causes an increase in [Ca²⁺]_cyt and that osmotic stress sharply decreases [Ca²⁺]_cyt in rice. The [pH]_cyt was differently shifted in the two rice cultivars in response to salt stress and may be coupled to different activities of the H⁺‐ATPases. The changes in vacuolar pH were correlated with the expressional analysis of rice vacuolar H⁺‐ATPase in these two rice cultivars.     

Serglycin constitutively secreted by myeloma plasma cells is a potent inhibitor of bone mineralization in vitro

Although the biological significance of proteoglycans (PGs) has previously been highlighted in multiple myeloma (MM), little is known about serglycin, which is a hematopoietic cell granule PG. In this study, we describe the expression and highly constitutive secretion of serglycin in several MM cell lines. Serglycin messenger RNA was detected in six MM cell lines. PGs were purified from conditioned medium of four MM cell lines, and serglycin substituted with 4-sulfated chondroitin sulfate was identified as the predominant PG. Flow cytometry and confocal microscopy showed that serglycin was also present intracellularly and on the cell surface, and attachment to the cell surface was at least in part dependent on intact glycosaminoglycan side chains. Immunohistochemical staining of bone marrow biopsies showed the presence of serglycin both in benign and malignant plasma cells. Immunoblotting in bone marrow aspirates from a limited number of patients with newly diagnosed MM revealed highly increased levels of serglycin in 30% of the cases. Serglycin isolated from myeloma plasma cells was found to influence the bone mineralization process through inhibition of the crystal growth rate of hydroxyapatite. This rate reduction was attributed to adsorption and further blocking of the active growth sites on the crystal surface. The apparent order of the crystallization reaction was found to be n=2, suggesting a surface diffusion-controlled spiral growth mechanism. Our findings suggest that serglycin release is a constitutive process, which may be of fundamental biological importance in the study of MM.     

Chymotryptic heavy meromyosin from gizzard myosin: a proteolytic fragment with the regulatory properties of the intact myosin.

Arginine deiminase (EC 3.5.3.6) from $\text{Mycoplasma}\text{arthritidis}$ is a dimeric enzyme. Velocity centrifugation in 6 M guanidine HCl and peptide mapping of the BrCN fragments suggest that the subunits are identical. The reaction of one out of four sulfhydryl groups with 0.3 mM 5,5′-dithiobis-(2-nitrobenzoic acid) has a half-life of about 30 min in 2 M guanidine HCl at 15°, pH 8. The enzyme is irreversibly inhibited by 1 mM formamidinium ion within 1 min. Inactivation by this affinity label is resolvable into two concurrent first-order reactions in the presence of guanidinium ion; the fraction of enzyme which reacts at the faster rate is about 50%. These results are interpreted as evidence for two catalytic subunits which differ in conformation.