Analysis of mRNA with microsomal fractionation using a SAGE-based DNA microarray system facilitates identification of the genes encoding secretory proteins

In the regulation of host defense responses such as inflammation and immunity, the secretory proteins, including membrane proteins, play central roles. Although many secretory proteins have been identified by using methods such as differential display, random screening, or the signal sequence trap method, each method suffers from poor reproducibility, low sensitivity, or time-consuming or laborious work. Therefore, the strategy for facilitating the selection of the genes encoding the secretory proteins is desired. In this paper, we describe a system for isolating the genes encoding secretory proteins by analyzing mRNAs with microsomal fractionation on serial analysis of gene expression (SAGE)-based DNA microarray system. This system succeeded in discriminating the genes encoding secretory proteins from ones encoding nonsecretory proteins with 80% accuracy. We applied this system to human T lymphocytes. As a result, we were able to identify the genes that are not only encoding secretory proteins but also expressing selectively in a specific subset of T lymphocytes. The SAGE-based DNA microarray system is a promising system to identify the genes encoding specific secretory proteins.     

A histological evaluation for guided bone regeneration induced by a collagenous membrane

This study was designed to evaluate the histological changes during ossification and cellular events including osteogenic differentiation responding to collagenous bioresorbable membranes utilized for GBR. Standardized artificial bony defects were prepared at rat maxillae, and covered with a collagenous bioresorbable membrane. These animals were sacrificed at 1, 2, 3 and 4 weeks after the GBR-operation. The paraffin sections were subject to tartrate resistant acid phosphatase (TRAP) enzyme histochemistry and immunohistochemistry for alkaline phosphatase (ALP), osteopontin (OP) and osteocalcin (OC). In the first week of the experimental group, woven bone with ALP-positive osteoblasts occupied the lower half of the cavity. The collagenous membrane included numerous ALP-negative cells and OP-immunoreactive extracellular matrices. At 2 weeks, the ALP-, OP- and OC-immunoreactivity came to be recognizable in the region of collagenous membrane. Since ALP-negative soft tissue separated the collagenous membrane and the new bone originating from the cavity bottom, the collagenous membrane appeared to induce osteogenesis in situ. At 3 weeks, numerous collagen fibers of the membrane were embedded in the adjacent bone matrix. At 4 weeks, the membrane-associated and the cavity-derived bones had completely integrated, showing the same height of the periosteal ridge as the surrounding alveolar bones. The collagen fibers of a GBR-membrane appear to participate in osteogenic differentiation.     

Anionic copolymerization of β-lactones in correlation with the mode of fission. studies on the syntheses of aliphatic polyesters. XI

The copolymerization behavior of β-propiolactone (PL) and α.α-bischloromethyl β-propiolactone (α-BCPL) is correlated with the mode of fission. With sodium acetate as catalyst, the formation of β-acetoxy propionic acid at the initiation reaction was observed and α-BCPL was more than five times as reactive as PL in copolymerization. Thus propagation by carboxylate anion through alkyl-oxygen fission was presumed in the case of pyridine and the acetate of Na, Li, and Mg as catalyst. On the other hand, the formation of ethyl β-hydroxy propionate and the decreased reactivity of α-BCPL in copolymerization was observed with magnesium and aluminum ethoxide as catalyst. In the case of the ethoxide of Na, Li, and Al, and Et₂Mg or Et₃Al as catalyst, propagation by alkoxide anions through acyl-oxygen fission of the coordinated lactones was presumed.     

Polymerization of methylmethacrylate initiated by hydrogenation catalysts

RANEY metals (Ni, Fe, Co), URUSHIBARA metals (Ni, Co) and ULIMANN cu initiate the polymerization of methylmethacrylate. Diamines, diols or organic halides enhance their reactivity. In the presence of CCl₄, the polymerization rate (R_p) is proportional to the square root of the amount of both metals and CCl₄, and to the first power of the monomer concentration, but R_p becomes constant and independent of the concentration of CCl₄ when there is a great excess of CCl₄. We concluded that this polymerization follows the free radical mechanism and the participation of a zero-valent metal atom in the initiation reaction including complex formation between metal and CCL₄ is suggested.     

Changes in the nuclear uptake and retention of 3H-estrogen in gonadotrophs and lactotrophs as a function of age

A quantitative autoradiographic immunocytochemical study was performed in which the nuclear uptake and retention of 3H-estradiol (3H-E2) by luteinizing hormone (LH) and prolactin (PRL) cells was examined in 19-21-year-old baboons. 3H-E2 concentrating cells were found in all of the three lobes of the pituitary in varying percentages (38.7%, pars distalis; 17.1%, pars intermedia; 6.3%, pars nervosa). Approximately 80% of PRL cells and nearly 100% of LH cells were labeled. A count of the number of silver grains over nuclei revealed a marked variation of the accumulation of 3H-E2 by LH cells and to a lesser extent in PRL cells. These results suggest functional heterogeneity among LH and PRL cells. The present results are discussed in relation to the physiological state of old animals.     

The determination of serum and urine myoglobin

We have reviewed the current methods of determining serum and urine myoglobin. Qualitative determination of urine myoglobin has been possible for over 20 years, but the quantitative determination is a recent development. Between 1963 (spectrophotometry) and 1978 (radioimmunoassay, RIA), efforts have been made to develop a more sensitive determination method. The RIA method thus developed has a sensitivity as high as 1 ng/ml. Such methods are valuable in the diagnosis of acute myocardial infarction. Recently, simple, rapid, and non-isotopic method such as enzyme immunoassay (EIA) and latex agglutination test have been developed and made rapid determination possible. We applied the "Mb-latex [Seiken]", one such latex agglutination method to the TBA-30 R single-multi type automated analyzer. We obtained good results in the fundamental study such as in precision and accuracy. The reference ranges obtained using serum and urine samples corresponded to the values reported by others with the exception of the lower limit of the ranges. Proficiency testing data using the same reagents and same samples at five laboratories revealed some systematic biases. The reasons for such interlaboratory differences is not clear. However, these systematic biases may be due to the type of equipment, the type of calibration procedure and others including reagents and standard solutions. We are continuing our effort to improve the reliability of interlaboratory data.     

VDE-initiated intein homing in Saccharomyces cerevisiae proceeds in a meiotic recombination-like manner

BACKGROUND: Inteins and group I introns found in prokaryotic and eukaryotic organisms occasionally behave as mobile genetic elements. During meiosis of the yeast Saccharomyces cerevisiae, the site-specific endonuclease encoded by VMA1 intein, VDE, triggers a single double-strand break (DSB) at an inteinless allele, leading to VMA1 intein homing. Besides the accumulating information on the in vitro activity of VDE, very little has been known about the molecular mechanism of intein homing in yeast nucleus. RESULTS: We developed an assay to detect the product of VMA1 intein homing in yeast genome. We analysed mutant phenotypes of RecA homologs, Rad51p and Dmc1p, and their interacting proteins, Rad54p and Tid1p, and found that they all play critical roles in intein inheritance. The absence of DSB end processing proteins, Sae2p and those in the Mre11-Rad50-Xrs2 complex, also causes partial reduction in homing efficiency. As with meiotic recombination, crossover events are frequently observed during intein homing. We also observed that the absence of premeiotic DNA replication caused by hydroxyurea (HU) or clb5delta clb6delta mutation reduces VDE-mediated DSBs. CONCLUSION: The repairing system working in intein homing shares molecular machinery with meiotic recombination induced by Spo11p. Moreover, like Spo11p-induced DNA cleavage, premeiotic DNA replication is a prerequisite for a VDE-induced DSB. VMA1 intein thus utilizes several host factors involved in meiotic and recombinational processes to spread its genetic information and guarantee its progeny through establishment of a parasitic relationship with the organism.     

A digest of the Evidence-Based Clinical Practice Guideline for Nephrotic Syndrome 2020

Clinical and Experimental Nephrology -     

Measurement of oil mists by a portable sampler and subjective symptoms among machine workers: a field investigation of machine workshops

Machine oils are widely used in metal processing and many workers are exposed to oil mists in the work environment. Some investigators have pointed out such health hazards due to prolonged exposure to oil mist as respiratory disorders, dermatitis, and possible carcinogenesis. In Japan, a permissible limit of oil mists of 3 mg/m3 was recommended in 1977 by the Japan Association of Industrial Health (JAIH) mainly based on the hazardous effects on the respiratory system. After this recommendation was made, only a few studies have been made on the measurement of oil mists and on the health effects in machine workers to determine whether the permissible limit is justifiable or not. In the present study, the levels of oil mists in the air of machine workshops were measured together with personal exposure levels by personal samplers. Oil mists were collected by a sampler head with 2 stages which enabled differentiation of distribution of particulates between larger than 10 microns and 2-10 microns in size. Oil components collected on the stainless steel stages were washed out by sonication in CCl4 solution and measured by an oil meter with infra-red spectrometry against a standard solution of heavy oil class B according to procedures reported elsewhere by the authors. Questionnaire surveys were also conducted on 308 male machine workers composed of 221 workers exposed to oil mists and 87 nonexposed controls. The questions were composed of five items about air quality in the work environment and 18 subjective symptoms during work and daily lives. The symptoms included nasopharyngeal, muco-dermal, gastrointestinal and neuro-muscular symptoms. Statistical analysis was made by the Manthel-Haenszel method for a comparison of "yes" rates of complaints between the exposed and the non-exposed by adjusting the underlying confounding factor of age distribution. 1. The levels of oil mists measured here ranged from 94 to 813 micrograms/m3 in the ambient air and from 107 to 483 micrograms/m3 of personal exposure. There was no obvious difference between the level in the ambient air and that of personal exposure. All these measured levels were under the permissible limit (3 mg/m3) recommended by JAIH in 1977. 2. The observed distribution of particulate size of oil mists from 2 to 10 microns referred to as respirable size was 32.8 +/- 16.1% generated from the grinding machines using water-soluble type of machine oils and 50.0 +/- 12.4% from those using insoluble type.(ABSTRACT TRUNCATED AT 400 WORDS)