Masaya Kitamura

Rat testis P-45017α cDNA: the deduced amino acid sequence, expression and secondary structural configuration

Summary A complete amino acid sequence for rat testis P-45017α was deduced from nucleotide analysis of a cDNA clone isolated from a rat Leydig cell cDNA library. This DNA clone, containing initiation and termination codons and a polyA tail, translated a polypeptide in COS-1 cells that expressed both 17α-hydroxylase and 17,20 lyase activities. It exhibited significant similarity to the nucleotide and deduced amino acid sequences of the bovine and human cytochrome P-45017α, particularly with respect to the highly conserved regions and secondary structure. The P-45017α appears to be anchored to the membrane of the endoplasmic reticulum through two transmembrane regions, specifically the N terminal insertion peptide and the stop-transfer sequence. Hydropathic analysis indicates that the remainder of the C terminus is associated with the membrane through four hydrophobic clefts, including the putative steroid binding site.

Chromosomal variants among 1790 infertile men

Abstract Background: The largest cytogenetic survey involving infertile men was undertaken to clarify whether chromosomal abnormalities, including autosomal abnormalities, affect semen qualities.

Prostate stromal cell-derived hepatocyte growth factor induces invasion of prostate cancer cell line DU145 through tumor-stromal interaction

In prostate cancer, several growth factors derived from stromal cells regulate tumor cell growth. Hepatocyte growth factor (HGF) possesses biological activities that promote cancer proliferation and invasion through tumor‐stromal interaction. We examined how prostate stromal cell‐derived HGF affects invasion of prostate cancer cells through this interaction.

REGULATION OF INVASIVE POTENTIAL OF HUMAN PROSTATE CANCER CELL LINES BY HEPATOCYTE GROWTH FACTOR

Background: The growth and progression of prostate cancer depends on the stromal‐epithelial interaction which is under paracrine control. Hepatocyte growth factor (HGF), produced by mesenchymal cells, is a multifunctional growth factor stimulating the movement and growth of epithelial cells including cancer cells. We therefore assessed the relationship between the invasive potential of prostate cancer and HGF in vitro.

Ejaculated spermatozoa in patients with non-mosaic Klinefelter's syndrome.

Background : Non‐mosaic Klinefelter patients are generally azoospermic and there is no therapy to improve the spermatogenesis. Some patients have a few spermatozoa in their ejaculates, which can be used for intracytoplasmic sperm injection (ICSI), but only a few cases resulting in a successful birth have been reported.

Rapid and simple determination of mycophenolic acid in human plasma by ion-pair RP-LC with fluorescence detection

Mycophenolic acid (MPA) is an immunosuppressive drug given as the prodrug of mycophenolate mofetil (MMF). In order to investigate the pharmacokinetics of MPA, a simple, specific, sensitive and reliable method has been established for the quantitative determination of MPA in plasma from renal transplant recipients. The method involves a single-step protein precipitation procedure and a specific determination by ion-pair reversed-phase high-performance liquid chromatography (HPLC) with fluorescence detection. Separation was achieved on a C18 column (150 x 4.6 mm, 5 microm) with a mobile phase composed of borate buffer (pH 10.0; 50 mM)--acetonitrile--tetrabutylammonium bromide (200 mM) (75:25:1, v/v/v). The fluorescence detector was set at 310 (excitation) and 430 nm (emission). Following protein precipitation with ice-cold acetonitrile, clear supernatants (50 microl) were injected into the HPLC system. The retention time of MPA was approximately 4.5 min. The HPLC run time was 8 min. The assay was linear in concentration range 0.2-20.0 microg/ml for MPA in human plasma. Precision of the assay in the concentration range examined was from 0.89 to 3.21% for the intra-assay run and from 3.01 to 4.35% for the inter-assay run. A limit of detection was 0.05 microg/ml at a signal-to-noise ratio of 3. This validated method was then applied to the determination of MPA concentrations in renal transplant recipients after oral administration of 0.75 g of MMF.

Rapid and simultaneous determination of mycophenolic acid and its glucuronide conjugate in human plasma by ion-pair reversed-phase high-performance liquid chromatography using isocratic elution

A high-performance liquid chromatographic method has been developed for the simultaneous determination of mycophenolic acid (MPA) and its glucuronide conjugate (MPAG) in human plasma. The method involves protein precipitation with acetonitrile, followed by ion-pair reversed-phase chromatography on C18 column, with a 40 mM tetrabutyl ammonium bromide (TBA)-acetonitrile (65:35, v/v) mobile phase. A 20-microl volume of clear supernatant was injected after centrifugation, and the eluent was monitored at 304 nm. No interference was found either with endogenous substances or with many concurrently used drugs, indicating a good selectivity for the procedure. Calibration curves were linear over a concentration range of 0.5-20.0 microg/ml for MPA and 5-200 microg/ml for MPAG. The accuracy of the method is good, that is, the relative error is below 5%. The intra- and inter-day reproducibility of the analytical method is adequate with relative statistical deviations of 6% or below. The limits of quantification for MPA and MPAG were lower than 0.5 and 5.0 microg/ml, respectively, using 50 microl of plasma. The method was used to determine the pharmacokinetic parameters of MPA and MPAG following oral administration in a patient with renal transplantation.

Possible association of infertility with sperm-specific abnormality of CD46.

Three infertile patients fulfilling normal or subnormal criteria on routine semen analysis showed abnormal sperm CD46 (membrane cofactor protein of complement) by SDS-PAGE/immunoblotting analysis using a panel of monoclonal antibodies. The sperm CD46 isoform has been reported to be associated with sperm-egg interaction. These three patients expressed normal CD46 isoforms on their lymphocytes and granulocytes. Sperm-specific abnormalities in these proteins thus parallel male infertility, suggesting a new category of infertility, probably due to aberrations in the molecules related to sperm-egg interaction.

Genomic analysis of idiopathic infertile patients with sperm-specific depletion of CD46.

Three infertile subjects with no expression of CD46 (membrane cofactor protein of complement) on their spermatozoa were found when screening 542 idiopathic male infertile patients. The sperm CD46 isoform was reported to be associated with the sperm-egg interaction, yet a ubiquitous expression of CD46 confers resistance to complement-mediated injury on host cells. All three patients expressed normal CD46 isoforms on their lymphocytes and granulocytes. Thus, the loss of CD46 is sperm-specific, probably due to testicular germ cell-specific regulation of CD46 production. Recently, a mechanism of the gene regulation of human CD46 was elucidated in which the silencer element of the 3′ UT and the promoter region of human CD46 gene partly participate. Here, we analyzed these regions of the CD46 gene in our 3 patients. We found no abnormality in 3′ and 5′ regions of the CD46 genome in the 3 patients. Thus, in these infertile patients sperm-specific depletion of CD46 is not governed by the so for identified regulators in the CD46 gene. Other unknown factors outside the known regulatory regions would play a role in the regulation of sperm-specific CD46 expression.

Effect of Renal Transplantation on Sexual Function

This investigation was conducted to determine whether renal transplantation can improve sexual function in male patients with chronic renal failure. The authors retrospectively studied 121 men undergoing renal transplantation who complained of any type or degree of sexual dysfunction pre-operatively. Sexual function was evaluated by questionnaire which included erectile, ejaculative, and orgasmic functions. Pre- and postoperative frequency of sexual intercourse was also recorded. Patient characteristics, laboratory data, and endocrinologic profiles were analyzed to identify factors that might influence sexual function. In patients with hormonal determinations, results essentially normalized after transplantation. However, only 43 patients (35.5%) reported improvement of overall sexual function after renal transplantation, while 34 (28.1%) reported worsening. Although frequency of sexual intercourse was unaffected by transplantation, 15 of 20 patients who had no intercourse before transplantation initiated intercourse afterward. These 15 patients all underwent transplantation before 40 years of age. Comparisons of variables by sexual function showed significant differences for type of immunosuppressive treatment, interval after renal transplantation, and serum concentration of hemoglobin A1c. It is concluded that renal transplantation cannot improve sexual function in allpatients, although hormonal profiles were largely normalized, and that renal transplantation should be encouraged at a younger age.