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Kazuhiko Hoshi

Publications and source records attributed to Kazuhiko Hoshi.

36 records · Page 2Linked to original sources
PubMed2004

Oocyte mitochondria: strategies to improve embryogenesis.

Mitochondria play a central role to provide ATP for fertilization and preimplantation embryo development in the ooplasm. The mitochondrial dysfunction of oocyte has been proposed as one of the causes of high levels of developmental retardation and arrest that occur in preimplantation embryos generated using Assisted Reproductive Technology. Cytoplasmic transfer (CT) from a donor to a recipient oocyte has been applied to infertility due to dysfunctional ooplasm, with resulting pregnancies and births. However, neither the efficacy nor safety of this procedure has been appropriately investigated. In order to improve embryogenesis, we observed the mitochondrial distribution in ooplasma under the several conditions using mitochondrial GFP-transgenic mice (mtGFP-tg mice) in which the mitochondria are visualized by GFP. In this report, we will present our research about the mitochondrial distribution in ooplasm during early embryogenesis and the fate of injected donor mitochondria after CT using mtGFP-tg mice. The mitochondria in ooplasm from the germinal vesicle stage to the morula stage were accumulated in the perinuclear region. The mitochondria of the mtGFP-tg mouse oocyte transferred into the wild type mouse embryo could be observed until the blastocysts stage, suggesting that the mtGFP-tg mice oocyte is very useful for visual observation of the mitochondrial distribution in the oocyte, and that the aberrant early developmental competences due to the oocyte mitochondrial dysfunction may be overcome by transferring the "normal" mitochondria.

Animals↗
PubMed2004

Milk consumption is a risk factor for prostate cancer: meta-analysis of case-control studies.

Prostate cancer has become the most common cancer among men in the United States. Although milk consumption is considered to be a risk factor in some epidemiological studies, the results are inconsistent. A meta-analysis method was conducted to estimate the combined odds ratio (OR) between milk consumption and prostate cancer from case-control studies published between 1984 and 2003 using commercial software (comprehensive meta-analysis). The combined OR was 1.68 (95% confidence interval = 1.34-2.12) in the 11 published case-control studies. The combined OR varied little by study stratification. Additionally, we evaluated the possible risk factors in milk for prostate cancer. In conclusion, we found a positive association between milk consumption and prostate cancer. The underlying mechanisms, including fat, calcium, hormones, and other factors, should be investigated further.

Age Factors↗
PubMed2004

Ubiquitin signals in the developing acrosome during spermatogenesis of rat testis: an immunoelectron microscopic study.

The localization of ubiquitin (UB) signals in the acrosomes of rat spermiogenic cells was investigated by immunoelectron microscopy using two anti-UB antibodies: UB1, reacting with ubiquitinated proteins and free UB; and FK1, recognizing polyubiquitinated proteins but not monoubiquitinated proteins or free UB. Labeling of UB by UB1 (UB1 signal) was detected in the acrosomes at any stage of differentiation. In step 1 spermatids, UB1 signals were detected on the cytoplasmic surface and in the matrix of transport vesicles located between the trans-Golgi network and the acrosome. Weak signals were detected in acrosomal granules within acrosome vesicles that had not yet attached to the nucleus. In step 4-5 spermatids, the acrosome vesicles had enlarged and attached to the nucleus. Strong gold labeling was noted in a narrow space between the outer acrosomal membrane and the developing acrosomal granule, where a dense fibrous material was observed on routine electron microscopy, whereas the acrosomal granule was weakly stained by UB1 antibody. In step 6-8 spermatids, UB1 signals were detected in the fibrous material that expanded laterally to form a narrow electronless dense zone between the acrosomal granule and the outer acrosomal membrane. Labeling in the acrosomal granule increased. In step 9-11 spermatids, UB1 signals were confined to the narrow zone from the tip of the head to the periphery of the ventral fin. The matrix of the acrosome was weakly stained. In epididymal sperm, UB1 labeling in the acrosome decreased without any pretreatment, whereas staining was noted in a spot in the neck region and in the dorsal fin after trypsin digestion. On the other hand, the staining pattern with FK1 was quite different from that with UB1. The trans-Golgi network was weakly stained but the cis-Golgi network was strongly stained. The dense fibrous material just beneath the outer membrane was never stained with FK1. The results suggest that UB on the surface of transport vesicles is involved in anterograde transport from the Golgi apparatus to the acrosome. The physiological role of UB in acrosomes is not clear. Two candidates for monoubiquitinated proteins in the acrosome, which have a UB-interacting motif, were found by cyber screening.

Acrosome↗
PubMed2003

Transient decrease in the CD45RA expression on T lymphocytes in neonates with fetal distress.

To determine the expression of CD45 isoforms on T lymphocytes in neonates with fetal distress and to evaluate its diagnostic accuracy, peripheral blood samples were examined in 53 neonates who were classified into one of three groups: group I: 'control' group (n = 23), group II: 'mild distress' group (n = 15), and group III: 'moderate distress' group (n = 15). The expression of CD3 (mean +/- SD 24.2 +/- 10.1%), CD4 (23.0 +/- 5.7%), and CD45RA (27.3 +/- 9.6%) on total lymphocytes and the expression of CD45RA on CD4+ T lymphocytes (13.7 +/- 4.7%) in group III were significantly lower than in the other two groups 0-3 days after birth. Sensitivity and specificity of the CD45RA expression on CD4+ T lymphocytes for discrimination of group III were calculated as 0.79 and 1.0, respectively, when the cutoff value was 22.7%. The low CD3, CD4, and CD45RA expression returned to normal levels 10 days and more after birth. There were no differences in the CD8 and CD45RO expression between the groups. We conclude that CD4+ T lymphocytes from neonates with fetal distress show a transient decrease in the CD45RA expression without an increase in the CD45RO expression, and, therefore, analysis of the CD45 isoform expression is useful for laboratory evaluation of fetal distress.

CD3 Complex↗
PubMed2003

Spatiotemporal changes of levels of a moonlighting protein, phospholipid hydroperoxide glutathione peroxidase, in subcellular compartments during spermatogenesis in the rat testis.

We studied temporal changes in the subcellular localization and levels of a moonlighting protein, phospholipid hydroperoxide glutathione peroxidase (PHGPx), in spermatogenic cells and mature sperm of the rat by immunofluorescence and immunoelectron microscopy. The PHGPx signals were detected in chromatoid bodies, clear nucleoplasm, mitochondria-associated material, mitochondrial aggregates, granulated bodies, and vesicles in residual bodies in addition to mitochondria, nuclei, and acrosomes as previously reported. Within mitochondria, PHGPx moved from the matrix to the outermost membrane region in step 19 spermatid, suggesting that this spatiotemporal change is synchronized with the functional change of PHGPx in mitochondria. In the nucleus, PHGPx was associated with electron-lucent spots and with the nuclear envelope, and PHGPx in the latter region increased after step 16. In early pachytene spermatids, PHGPx signals were noted in the nuclear material exhibiting a very similar density to chromatoid bodies and in the intermitochondrial cement, supporting the previous proposal that chromatoid bodies originate from the nucleus and intermitochondrial cement. The presence of PHGPx in such various compartments suggested versatile roles for this protein in spermatogenesis. Quantitative immunoelectron microscopic analysis also revealed dynamic changes in the labeling density of PHGPx in different subcellular compartments as follows: 1). Total cellular PHGPx rapidly increased after step 5 and reached a maximum at step 18; 2). mitochondrial labeling density increased after step 1 and achieved a maximum in steps 15-17; 3). nuclear labeling density suddenly increased in steps 12-14 to a maximum; 4). in cytoplasmic matrix, the density remained low in all steps; and 5). the labeling density in chromatoid bodies gradually decreased from pachytene spermatocytes to spermatids at step 18. These spatiotemporal changes in the level of PHGPx during the differentiation of spermatogenic cells to sperm infer that PHGPx plays a diverse and important biological role in spermatogenesis.

Animals↗
PubMed2003

Isoform/variant mRNAs for sex steroid hormone receptors in humans.

The open reading frames of human sex steroid hormone receptors (hSSHRs) are composed of eight exons. In addition, the presence of various exons - including 5'-untranslated exons, alternative coding exons and novel 'intronic' exons - has been demonstrated in the genes encoding hSSHRs. The isoform/variant hSSHR mRNAs generated from thes e exons can be tentatively classified into seven types. In type 1, different mRNAs are generated with the use of alternative transcription start sites. In type 2, one or more exons are skipped. In type 3, one or more exons are duplicated. In type 4, distinct mRNAs containing different 5'-untranslated exon(s) are synthesized. In type 5, distinct mRNAs possessing different coding exon(s) are generated. In type 6, mRNA is synthesized by intronic exons and coding exons 4/5-8. In type 7, mRNA with insertion of intronic exon(s) is generated. Here, we review the isoform/variant hSSHR mRNAs and the structure of the genes encoding them.

Alternative Splicing↗
PubMed2003

The relationship between carbohydrate intake and glucose tolerance in pregnant women.

OBJECTIVE: We verified whether a misdiagnosis of gestational diabetes mellitus can result in pregnant women when glucose tolerance has deteriorated after a low-carbohydrate meal, and tried to elucidate the mechanism behind the different outcome of the test. STUDY DESIGN: Twenty-seven pregnant women were given directions for their evening meal the day before each of two 75-g oral glucose tolerance tests (OGTT). The evening meal was either a low-carbohydrate meal (carbohydrate, 6.7%; Low), or a high-carbohydrate meal (carbohydrate, 85.7%; High). RESULTS: The OGTT showed that the glucose tolerance was significantly impaired after Low than after High, with a significant increase of fasting plasma non-esterified fatty acids (NEFA) level. Moreover, the insulinogenic index (I-I) after High significantly decreased than that after Low. CONCLUSIONS: The present data suggests that there is a risk of misdiagnosis of impaired glucose tolerance with only one intake of this extremely low-carbohydrate meal on the evening before testing. The decrease of insulin secretion and the activation of glucose-fatty acid cycle may be considered as the mechanism.

Adult↗
PubMed2003

The novel 5'-untranslated first exon, exon 0H, of the rat estrogen receptor beta gene.

The multiple untranslated first exons and promoters system has been reported to be involved in the tissue-specific expression of the estrogen receptor alpha (ERalpha) in humans and rats. However, a few reports are available concerning tissue-specific regulation of the expression of the estrogen receptor beta (ERbeta) gene. To investigate the mechanism regulating the expression of the rat ERbeta gene, we analyzed the structure of the 5'-untranslated region (UTR) of the rat testicular ERbeta mRNA using 5'-rapid amplification of the cDNA ends (5'-RACE) method. Sequence analysis revealed the presence of two isoforms of the ERbeta mRNA containing distinct 5'-UTRs. Although the 5'-UTR of one isoform of the messages was identical to the 5'-UTR of the previously reported ERbeta cDNA, the other isoform had a novel sequence in its 5'-UTR. Genomic analysis revealed that the 5'-UTRs of these two mRNA isoforms originated from two distinct untranslated first exons, the previously identified exon termed "exon 0N," and the novel exon we termed "exon 0H," both of which were spliced onto exon 1. We termed these isoforms of the messages containing the exon 0N and exon 0H, the ERbeta mRNA (0N-1) and ERbeta mRNA (0H-1), respectively. Furthermore, the distributions of these mRNA isoforms in several rat tissues were analyzed using the reverse transcription-polymerase chain reaction (RT-PCR) method. The distributions of the two mRNA isoforms differed; the ERbeta mRNA (0N-1) was widely distributed in the tissues examined, while expression of the ERbeta mRNA (0H-1) was restricted to a few tissues such as the anterior pituitary, amygdala, and some peripheral tissues. In conclusion, our findings indicate that the tissue-specific expression of the rat ERbeta gene is regulated, at least in part, by the multiple untranslated first exons system which consists of exon 0N and exon 0H.

5' Untranslated Regions↗
PubMed2003

Wilms tumor gene immunoreactivity in primary serous carcinomas of the fallopian tube, ovary, endometrium, and peritoneum.

Wilms tumor gene (WT-1) expression has been reported in many human cancers, including most ovarian and peritoneal serous carcinomas, but has not been studied in carcinomas of the fallopian tube. In this study, the authors evaluated the immunohistochemical expression of WT-1 in serous carcinomas of the fallopian tube and compared their reactivity with that of ovarian, peritoneal, and endometrial serous carcinomas. All primary serous carcinomas of the fallopian tube (13 cases), ovaries (25 cases), and peritoneum (3 cases) were reactive with the WT-1 antibody, whereas all five primary endometrial serous carcinomas were nonreactive. WT-1 reactivity in an unknown primary serous carcinoma is therefore suggestive of an extrauterine site. The marked difference in WT-1 staining raises the possibility of genetic differences between serous carcinomas arising in the endometrium compared with those arising in the ovaries, fallopian tubes, and peritoneum.

Adult↗
PubMed2003

Novel splicing events of untranslated first exons in human estrogen receptor alpha (ER alpha) gene.

In order to analyze the structures of the 5'-untranslated region of estrogen receptor alpha (ER alpha) mRNA in human uterine endometrium (Em), total RNA from Em was analyzed by 5'-rapid amplification of the cDNA ends method with antisense primer located on exon 1 of human ER alpha gene. Three isoforms of 5'-RACE clones were obtained: ER alpha mRNAs containing exon (A) (the upstream region of exon 1), exon C, and exons F-E2 (we adopted the nomenclature of 5'-untranslated exons of the Gannon group). The results imply that the major isoforms of ER alpha mRNA expressed in Em are these three isoforms. Moreover, reverse transcription-polymerase chain reaction (RT-PCR) analysis was carried out on Em, ovary (Ov) and liver (Li) mRNAs to detect the novel isoforms of ER alpha mRNA in these tissues, using sense primers located on exons (A), B, C, F, and E1, and antisense primer located on exon 1. As a result, in addition to the previously reported ER alpha mRNA isoforms containing exons (A), B, C, F-E2 and E1-E2 on exon 1, we identified two novel isoform mRNAs in which exons F and E1 were directly spliced onto exon 1. Differential distributions of these isoforms of ER alpha mRNAs in Em, Ov and Li were demonstrated by RT-PCR-Southern blot analysis. These results, together with the previous reports by others, indicate that there are at least ten isoforms of ER alpha mRNA containing different 5'-untranslated regions, exons (A), B, C, D, T1-T2, T1, F-E2, F, E1-E2 and E1, expressed in human, and that these are involved in tissue specific expression of the gene.

Base Sequence↗
PubMed2002

The novel exon, exon T, of the human progesterone receptor gene and the genomic organization of the gene.

Recently, we have cloned the novel isoform of the progesterone receptor (PR) cDNA (PR isoform S cDNA) from the human testicular cDNA library. The isoform S cDNA consists of the novel exon (termed the exon S of the PR gene) and the exons 4-8 of the PR gene. In order to investigate the existence of the other isoform of the human PR cDNA, the human testicular cDNA library was screened by the exons 4-8 corresponding sequence of the human PR cDNA in the present study. As a result, we have identified a novel isoform of the PR cDNA (termed the PR isoform T cDNA (PR-T cDNA)), which consisted of a previously unidentified 5'-sequence and the exons 4-8 of the PR gene. The structure of this isoform T cDNA is essentially similar to that of the isoform S cDNA. By the genomic cloning, the 5'-sequence of the PR isoform T mRNA was demonstrated to originate from a novel independent exon, exon T, which was located in the 5'-upstream region of the exon S.

Base Sequence↗
PubMed2002

The novel isoform of the estrogen receptor-alpha cDNA (ERalpha isoform S cDNA) in the human testis.

In order to clone the novel isoform of the cDNA for the human estrogen receptor-alpha (ERalpha), the human testicular cDNA library was screened by the exons 4-8 corresponding sequence of the human ERalpha cDNA. As a result, a novel isoform of the ERalpha cDNA (termed the ERalpha isoform S cDNA), which consists of a previously unidentified 5'-sequence and the exons 4-8 of the ERalpha gene, has been cloned. The structure of the ERalpha isoform S cDNA is essentially similar to that of the progesterone receptor (PR) isoform S cDNA that was identified in our recent report. Analysis of the genomic DNA revealed that the 5'-sequence of the ERalpha isoform S mRNA originated from a novel exon (termed the exon S). Moreover, the reverse transcription-polymerase chain reaction (RT-PCR) was carried out using the primers specific to the ERalpha isoform S mRNA on the total RNA from the human spermatozoon (Sp), liver (Li), uterine endometrium (Em) and myometrium (Mm). The ERalpha isoform S mRNA was detected in the uterine Em and Sp. Moreover, the molecular size of the ERalpha isoform S encoded by the ERalpha isoform S mRNA, which was analyzed by the transfection of the expression vector with ERalpha isoform S cDNA into the 293T cell, was approximately 39kDa. It was indicated that the one of the ATGs in the exon S could be used as the translation initiation codon. This is the first report on the ERalpha mRNA isoform that is not caused by exon-skipping or alternative utilization of the untranslated 5'-exons.

Base Sequence↗
PubMed2002

Cloning of the novel isoform of the estrogen receptor beta cDNA (ERbeta isoform M cDNA) from the human testicular cDNA library.

Our recent report has revealed the existence of the progesterone receptor (PR) isoform S, which consists of the novel PR exon S and exons 4-8 of the PR gene in the human testicular cDNA library. More recently, we have cloned the human estrogen receptor alpha (ERalpha) isoform S cDNA from the library. The ERalpha isoform S cDNA also contains the novel ERalpha exon S and exons 4-8 of the ERalpha cDNA. Based on these findings, we assumed that the novel isoform of cDNA like the PR- and ERalpha isoforms might exist in the human ER beta (ERbeta). In order to investigate this possibility, we have screened the human testicular cDNA library using the exons 4-8 corresponding sequence of the human ERbeta cDNA. Consequently, we have cloned a novel isoform of the ERbeta cDNA that consists of a previously unidentified 5'-sequence and the exons 5-8 of the ERbeta gene. We termed this isoform cDNA the "ERbeta isoform M cDNA". The 5'-sequence of the ERbeta isoform M cDNA was confirmed to be derived from a novel exon (termed the "exon M") by analysis of the genomic DNA. Moreover, we have analyzed the molecular size of the ERbeta isoform M encoded by the ERbeta isoform M mRNA by transient expression of the ERbeta isoform M cDNA in the 293T cell. The approximately 28 kDa protein, which was recognized by the anti-rat ERbeta antibody against the carboxyl-terminal region, was synthesized in the cells. Thus, we concluded that the ATG in the exon M could be used as the translation initiation codon. This report revealed for the first time the existence of the ERbeta mRNA isoform that is not caused by the skipping of one or more exons, by the alternative usage of the multiple exon 8s, nor by the alternative utilization of the untranslated 5'-exons located on the upstream region of the exon 1.

Alternative Splicing↗
PubMed2002

Novel isoforms of the mRNA for human female sex steroid hormone receptors.

In our recent reports, the novel isoform cDNAs of the ER alpha (ER alpha isoform S cDNA), ER beta (ER beta isoform M cDNA) and PR (PR isoform S and PR isoform T cDNAs) have been identified. These isoform cDNAs contained the previously unidentified 5'-sequences on exons 4-8 (ER alpha isoform S cDNA), exons 5-8 (ER beta isoform M cDNA) or exons 4-8 (PR isoform S and PR isoform T cDNAs). The genomic DNA analysis revealed that the 5'-sequences were derived from the novel independent exons, the ER alpha exon S, ER beta exon M, PR exon S and PR exon T, respectively. Furthermore, the existence of the novel variant mRNA, termed the i45 PR mRNA variant, with the insertion of the previously unidentified exons, termed the exons i45a and i45b, has been demonstrated by the reverse transcription-polymerase chain reaction on the RNA of the human uterine endometrium. From these results, we have concluded that the genes for the human female sex steroid hormone receptors contain the novel intronic exons, that the novel isoform mRNAs are transcribed using the intronic exon and exons 4-8 (or exons 5-8) of the gene, and that the novel variant mRNA is generated by the insertion of the intronic exons in the PR. In the present communication, our recent data along with others on the novel isoform/variant mRNAs for the human female sex steroid hormone receptors will be summarized.

5' Untranslated Regions↗
PubMed2002

Fertilization and embryo development in a mouse ICSI model using human and mouse sperm after immobilization in polyvinylpyrrolidone.

BACKGROUND: For human ICSI, sperm are normally immobilized immediately prior to injection. However, there are some situations when only sperm of questionable viability are available. There are few evaluations of fertilization or developmental problems in human or animal models using sperm having known intervals between immobilization and injection. METHODS: Immobilized human sperm were maintained for 1-24 h in 10% polyvinylpyrrolidone (PVP) before injection into mouse oocytes. Mouse sperm heads were similarly maintained in either PVP or a high potassium-containing 'nucleus isolation medium' (NIM) before ICSI and embryo development to the blastocyst stage. RESULTS: Immobilized human sperm activated mouse oocytes comparably to controls even 24 h after immobilization. However, mouse sperm heads showed a decrease in activating ability 6 h after isolation, either in PVP or NIM. A significant reduction in blastocyst development occurred if mouse sperm heads were maintained for even 1 h in PVP. After 6 h, no blastocysts formed, with arrest occurring at the morula stage. NIM provided partial protection for up to 3 h. CONCLUSIONS: Immobilized human sperm maintained oocyte activating activity for 24 h. However, mouse sperm are susceptible to alterations that affect both fertilization and development.

Animals↗
PubMed2002

Progesterone receptor mRNA variant containing novel exon insertions between exon 4 and exon 5 in human uterine endometrium.

The presence of human progesterone receptor (PR) mRNA variants has been demonstrated in uterine endometrium and breast tissues as well as in cancer cells of these tissues. While exon deletions by the alternative splicing in these variants have been reported, there are few reports available on the PR mRNA variants with exon insertion. In the present study, we attempted to detect a PR mRNA variant containing the exon insertions in normal uterine endometrium. Endometrial tissues were subjected to reverse transcription-polymerase chain reaction (RT-PCR) with PCR primers which were located in exons 3 and 8. Analysis of the RT-PCR products revealed the presence of a novel PR mRNA variant which contained a 232 bp inserted nucleotide sequence between exons 4 and 5. We termed this transcript the "i45 PR mRNA variant". Genomic analysis indicated that the inserted sequence was derived from two novel independent exons of 123 bp and 109 bp, termed "exon i45a" and "exon i45b", respectively, which are located between exons 4 and 5 of the human PR gene. The i45 PR mRNA variant was further detected in uterine endometrial cancer tissues as well as in the normal uterine endometrium. These results demonstrate the presence of a novel PR mRNA variant with exon insertions in the human tissue for the first time. The i45 PR variant protein, possibly transcribed from this i45 PR mRNA variant, may play physiological and/or pathological roles in the human uterine endometrium.

Base Sequence↗
PubMed2002

Relationship between sperm mitochondrial membrane potential, sperm motility, and fertility potential.

AIM: To analyze the relationship between sperm mitochondrial membrane potential and sperm motility parameters by means of a computer-assisted sperm analyzer (CASA) and in-vitro fertilization rate(%FR). METHODS: Semen samples were obtained from 26 men undergoing in vitro fertilization-embryo transfer (IVF-ET). Informed consent was obtained from all men prior to the study. Samples were prepared using wash and swim-up method in HEPES-HTF medium. The sperm motility (%MOT), progressive motility (%PMOT), average path velocity (VAP) microm/s), straight line velocity (VSL) (micro m/s), curvilinear velocity (VCL) (microm/s) and %hyperactivated sperm (%HA), and the %FR were assessed. The samples were incubated in the presence of 2.0 mciromol/L of 5,5',6,6'-tetra-chloro-1,1',3,3'-tetraethylbenzimidazolyl-carbocyanine iodide (JC-1) for 30 min at 37 degrees C in air and washed in PBS before flow cytometry (FACSCalibur: Becton Dickinson) analysis. The mitochondrial probe JC-1 was used to identify the mitochondrial membrane potential. The sperm was divided into three populations according to the fluorescence pattern as follows: the high mitochondrial membrane potential group (n=8), the moderate group (n=5), and the low group (n=13). Statistical analysis was performed using unpaired t-test. RESULTS: Significant differences were found between the high and the low groups in %MOT (91.1+/-8.5 vs 63.0+/-32.7, mean+/-SD), VAP (73.0+/-14.2 vs 52.1+/-12.5), VCL (127.0+/-28.1 vs 87.0+/-22.6), %HA (27.3+/-23.6 vs 7.2+/-9.0) and %FR [73.2 (48/56) vs 59.0 (69/117)]. No significant differences were found in other CASA parameters. CONCLUSION: When the sperm mitochondrial membrane potential increases, sperm motility parameters and fertility potential will also increase. The JC-1 dye method is useful to predict sperm fertility potential.

Embryo Transfer↗