Effects of renal function on the pharmacokinetics and pharmacodynamics of prophylactic cefazolin in cardiothoracic surgery.

The purpose of this investigation was to study the effects of renal function on the pharmacokinetics and pharmacodynamics (PK-PD) of free cefazolin administered prophylactically in cardiothoracic surgery. Patients received an initial 2-g dose of cefazolin, followed by 1-g doses 6, 12, 18 and 24 h after the first dose. In patients who underwent cardiopulmonary bypass, 1 g was added to the priming solution. In 35 patients with a normal estimated creatinine clearance (CLcr) ≥50 ml/min, a free cefazolin concentration <4 µg/ml was observed in 11.4, 5.7 and 54.3% of patients before the second dose, at the end and 24 h after operation, respectively. In contrast, only 7.4% of 27 patients with CLcr <49 ml/min had a free cefazolin concentration <4 µg/ml 24 h after the operation. There was a high negative correlation between CLcr and time above the target minimal inhibitory concentration (MIC) when the CLcr was <50 ml/min (r(2) = 0.807), and no correlation when the CLcr was ≥50 ml/min. Renal function has a significant impact on the PK-PD of prophylactic cefazolin in cardiothoracic surgery. The postoperative drug dosing intervals should be <6 h in order to achieve a 100% time above the MIC in patients with CLcr ≥ 50 ml/min.

Pregnancy and delivery following sonohysterographic lysis to treat recurrence after hysteroscopic lysis of severe intrauterine adhesions: a case report.

OBJECTIVE: To report a patient with sterility secondary to severe intrauterine adhesions who underwent sonohysterographic (SHG) lysis for recurrent adhesions following hysteroscopic lysis, and achieved tubal patency and natural pregnancy leading to term vaginal delivery. DESIGN: Case report. SETTING: National Hospital Organization Kyoto Medical Center, Kyoto, Japan. PATIENT: A patient with hypomenorrhea and sterility due to postpartum severe intrauterine adhesions. INTERVENTIONS: Operative hysteroscopy was performed for the severe intrauterine adhesions, and SHG lysis was performed for each of the recurrent adhesions that had occurred four times. RESULTS: SHG lysis improved the hypomenorrhea and restored the patency of the occluded fallopian tube. The patient became pregnant, and vaginally delivered a full-term infant. CONCLUSION: This approach may be an option if recurrent adhesions following hysteroscopic lysis occur.

Apoptosis in human endometrium and endometriosis.

Apoptosis plays a critical role in maintaining tissue homeostasis and represents a normal function to eliminate excess or dysfunctional cells. Accumulated evidence suggests that apoptosis helps to maintain cellular homeostasis during the menstrual cycle by eliminating senescent cells from the functional layer of the uterine endometrium during the late secretory and menstrual phase of the cycle. The BCL-2 family and Fas/FasL system have been extensively studied in human endometrium and endometriotic tissues. Eutopic endometrium from women with endometriosis reportedly has some fundamental differences compared with normal endometrium of women without endometriosis. The differences could contribute to the survival of regurgitating endometrial cells into the peritoneal cavity and the development of endometriosis. One mechanism that recently gained a lot of interest is the finding that apoptosis appeared in eutopic and ectopic endometrium of patients with endometriosis. This study is a current review of the literature focused on the physiological role of apoptosis in normal endometrium and the alterations in regulation of apoptosis in eutopic and ectopic endometrium from women with endometriosis. Similarities in characteristics of endometriosis at a molecular level with gynaecological tumours are also discussed. Finally, the role of apoptosis in the treatment of endometriosis is reviewed to link the basic research findings into clinical applications.

The Gly367Arg mutation in the myocilin gene causes adult-onset primary open-angle glaucoma.

PURPOSE: To present the phenotype of a family whose members showed the Gly367Arg mutation in the myocilin gene and developed primary open-angle glaucoma (POAG). METHODS: The proband developed POAG when she was 45 years old. Examination of the myocilin gene revealed that the patient had a mutation causing amino-acid change (Gly367Arg) in the myocilin gene. The available family members were given clinical and genetic examinations. RESULTS: Eight members of this family carried the same mutation. The age of disease onset of POAG in these patients with the mutation averaged 36.7 years. Four young members with the mutation, with an average age of 20.8 years, had not yet developed POAG. CONCLUSION: The Gly367Arg mutation of the myocilin gene in the pedigree causes the development of POAG in adulthood.

Molecular cloning of the bovine MYOC and induction of its expression in trabecular meshwork cells.

PURPOSE: Myocilin gene (MYOC) was identified as one of the disease-causing genes of primary open-angle glaucoma. This study was conducted to establish a system for the investigation of the biological role of MYOC in vitro by using bovine eyes, which are easy to obtain and have been widely used to examine the aqueous outflow system. The cDNA sequence of the bovine MYOC was determined and its expression in bovine eyes was examined with a quantitative polymerase chain reaction (PCR) assay. METHODS: Bovine MYOC cDNA was obtained from cultured bovine trabecular meshwork cells, and part of its sequence was determined using a primer pair designed based on the known sequence of the human MYOC gene. The 3' and 5' ends of this sequence were determined using the method of 3' and 5' rapid amplification of cDNA ends. The induction of the MYOC gene in cultured bovine trabecular meshwork cells after exposure to dexamethasone was quantitatively examined with real-time quantitative PCR using a probe designed according to the sequence of the determined bovine MYOC gene. RESULTS: Bovine MYOC protein was composed of 490 amino acids, which was 81.6% identical with that of human MYOC protein. Most of the amino acid residues of which mutation was reported to cause glaucoma were conserved in the bovine MYOC protein. After 2 weeks of treatment with 500 nM dexamethasone, expression of bovine MYOC mRNA was amplified 14-fold (14.1+/-5.1-fold, mean +/- SEM) measured by real-time quantitative PCR. CONCLUSIONS: The cDNA sequence of the bovine MYOC gene had a high degree of similarity to that of the human MYOC gene. Investigation of the function of bovine MYOC may contribute to identifying the role of MYOC protein in the aqueous outflow system.

Keratinocyte growth factor in the promotion of human chorionic gonadotropin production in human choriocarcinoma cells.

OBJECTIVE: Several growth factors and cytokines appear to participate in the proliferation or differentiation of trophoblast cells. The purpose of this study was to investigate the extent to which keratinocyte growth factor participates in the development of human embryonic and trophoblast cells at the maternal-fetal interface. STUDY DESIGN: The reverse transcriptase-polymerase chain reaction method was used to determine the gene expression of keratinocyte growth factor and keratinocyte growth factor receptor in human choriocarcinoma cells (BeWo), human teratocarcinoma cells (PA-1), and human endometrial stromal cells. We also examined the effects of keratinocyte growth factor on cell proliferation and production of human chorionic gonadotropin in BeWo and PA-1 cells. RESULTS: Keratinocyte growth factor gene was expressed in all cell types. The expression was pronounced in stromal cells of the endometrium collected during the secretory phase and early pregnancy. The keratinocyte growth factor expression was also enhanced in the differentiated BeWo cells. The expression of keratinocyte growth factor receptor gene was observed only in the BeWo cells. The addition of keratinocyte growth factor to the medium did not affect cell proliferation of the BeWo and PA-1 cells. On the other hand, keratinocyte growth factor (100 ng/mL) significantly enhanced human chorionic gonadotropin production in the BeWo cells. Stimulatory action of keratinocyte growth factor on human chorionic gonadotropin production in the BeWo cells was markedly enhanced after forskolin-induced differentiation. CONCLUSIONS: We conclude that keratinocyte growth factor may play an important role in promotion of human chorionic gonadotropin production in the trophoblast cells.

Altered gene expression and secretion of interleukin-6 in stromal cells derived from endometriotic tissues.

OBJECTIVE: To compare the expression of interleukin-6 (IL-6) in endometrial and endometriotic cells. DESIGN: Prospective study. SETTING: Department of Obstetrics and Gynecology, Tottori University Hospital, Yonago, Japan. PATIENT(S): Twenty patients who underwent either hysterectomy or laparoscopic surgery. INTERVENTION(S): Endometrial and endometriotic stromal cells were obtained from normal endometrium and from chocolate cyst linings of the ovary. Peritoneal macrophages were isolated from peritoneal fluids. Cells were cultured in the presence or absence of tumor necrosis factor-alpha. MAIN OUTCOME MEASURE(S): Gene expression of IL-6 was examined by Northern blot analysis. Interleukin-6 protein production was examined by immunocytochemical staining and ELISA. RESULT(S): A single IL-6 messenger RNA band of approximately 1.3 kilobases was detected in endometriotic stromal cells. Tumor necrosis factor-alpha increased the expression of IL-6 messenger RNA in endometriotic cells in a dose-dependent manner. In endometrial stromal cells, IL-6 messenger RNA signals were much weaker. Endometriotic stromal cells produced significantly larger amounts of IL-6 compared with endometrial stromal cells under basal conditions and after stimulation with tumor necrosis factor-alpha. Interleukin-6 protein was detected in cells isolated from endometriotic tissues by immunocytochemical staining. Interleukin-6 production by cultured macrophages from patients with endometriosis and endometriotic stromal cells was comparable. CONCLUSION(S): Altered gene expression and protein secretion of IL-6 in patients with endometriosis may contribute to the pathogenesis of the disease and/or to endometriosis-associated infertility.

Immunohistochemical localization of MYOC/TIGR protein in the trabecular tissue of normal and glaucomatous eyes.

PURPOSE: To examine immunohistochemically the localization of myocilin/trabecular meshwork inducible glucocorticoid response (MYOC/TIGR) protein in the glaucomatous and normal trabecular meshworks. METHODS: Trabecular tissues were used from one eye with late-onset goniodysgenetic glaucoma, three with primary open angle glaucoma (one of which had the MYOC/TIGR gene mutation), two with exfoliation glaucoma and one without glaucoma. For light microscopic immunohistochemistry, frozen sections were stained by the avidin-biotin complex method using anti-MYOC/TIGR polyclonal antibody. For electron microscopic immunohistochemistry, the pre-embedding method using the same antibody was performed. Double immunostaining using both anti-MYOC/TIGR and anti-type VI collagen antibodies was done by the immunofluorescence method. RESULTS: With light microscopy, immunoreactivity was seen in the whole trabecular meshwork of each of the specimens. No notable differences were detected in staining among the types of glaucoma, or between the eyes with and those without the gene mutation. Under electron microscopy, immunoreaction products were observed not only in the cytoplasm of the trabecular cells but also in the extracellular matrix, where staining was associated with the long-spacing collagen, fine granular materials and possibly microfibrils. With double immunohistochemistry, MYOC/TIGR was colocalized with type VI collagen in the trabecular meshwork. CONCLUSIONS: In glaucomatous and normal trabecular meshworks, the MYOC/TIGR protein is distributed in the extracellular matrix colocalizing with type VI collagen.

Laparoscopic surgery for endometriosis: a long-term follow-up.

OBJECTIVE: To investigate if complete resolution of endometriosis by laparoscopic surgery is beneficial to postoperative fecundity, dysmenorrhea and dyspareunia. DESIGN: An observational comparative study on the outcome of laparoscopic surgery. PATIENTS: Laparoscopically-treated symptomatic women with endometriosis (total n = 236); complete (n = 185) and incomplete (n = 51) surgery groups. MEASUREMENTS: Postoperative fecundity and symptom reduction. RESULTS: With whole populations, no surgical completeness-related difference was observed in cumulative pregnancy rates during the postoperative days 0-400 (cycle fecundity rate = 0.0319). Further accumulation of pregnant cases was followed in the complete surgery group (final cumulative pregnancy rate = 80%), but not in the counterpart group (p = 0.003). The similar result was obtained when only r-AFS classification stages III and IV were compared (p = 0.007). No r-AFS stage-related difference was observed in cumulative pregnancy rates when only patients of complete surgery were selected for comparison. The surgery reduced dysmenorrhea (84.7%) and dyspareunia (80.0%). CONCLUSIONS: Laparoscopic conservative surgery for endometriosis, especially when it is complete, increases fecundity and reduces disease-related symptoms, such as dysmenorrhea and dyspareunia.

The potential role of stem cell factor and its receptor c-kit in the mouse blastocyst implantation.

Embryo implantation is a complex process that requires the interaction of embryo and endometrium. Several growth factors and cytokines appear to be involved in this process. Stem cell factor (SCF) and its receptor c-kit regulate the proliferation and survival of germ cells and play an important role in follicular development. However, little information is available on the role of SCF and c-kit in the process of blastocyst implantation. In the present study, we examined the expression of SCF and c-kit mRNA in mouse embryos and in the stromal and epithelial cells of the uterine endometrium by reverse transcription-polymerase chain reaction (RT-PCR). SCF mRNA was expressed in the spreading blastocysts and endometrial cells, with especially strong expression occurring in the stromal cells. Expression of c-kit mRNA was detected in the blastocysts and spreading blastocysts, as well as in the endometrial cells. By immunocytochemical studies, staining for c-kit protein was observed in the in-vitro spreading trophoblasts. We found that 50-100 ng/ml SCF significantly promoted the expansion of the surface area of the spreading blastocysts (P < 0.01). These results are consistent with the hypothesis that SCF derived from endometrial cells and the implanting embryo exerts paracrine and/or autocrine action on the process of implantation by stimulating trophoblast outgrowth through its receptor c-kit.

Augmentation of local antitumor immunity in the liver by tumor vaccine modified to secrete murine interleukin 12.

Minimal residual lesions have been a major problem in surgical management of cancer. We transfected M5076 with murine IL-12 gene by a retroviral vector, established a stable transfectant secreting IL-12 and investigated its antitumor effects on a spontaneous liver metastasis murine model of M5076 reticulum cell sarcoma. Subcutaneous vaccination of the irradiated transfectant into the remote skin following the amputation of the tumor-bearing limb improved survival when compared with the vaccination of irradiated parental cells (control). Cytotoxic activities against parental M5076 were significantly stronger in the hepatic lymphocytes from the mice vaccinated with the IL-12 transfectant than those from the control. IFN-gamma production of hepatic lymphocytes when they were cocultured with the parental cells was significantly augmented in mice vaccinated with the IL-12 transfectant compared with the control. On the other hand, both cytotoxic activity and IFN-gamma production of spleen cells in the M5076-vaccinated and transfectant-vaccinated mice were at similar levels. Immunophenotypic analysis revealed the selective increase of CD3+NK1+ population in the liver from the transfectant-vaccinated mice. These results suggest that tumor vaccines genetically modified to secrete IL-12 continuously at a relatively low level preferentially augment local antitumor activity in the liver rather than systemic immune responses. This strategy warrants further investigation as an adjuvant modality in the management of postoperative residual tumors.

Unilateral papilledema after bone marrow transplantation.

We describe a patient who developed unilateral papilledema after allogeneic BMT. This is a rare manifestation of pseudotumor cerebri, which results from elevated intracranial pressure caused by cyclosporin A. The papilledema usually involves the fundi bilaterally, but unilateral involvement has been described. Congenital anomalies, compression and adhesion of the optic nerve sheath are its causes. In this patient, the right optic fundus was spared although leukemic infiltration was present on this side and high-dose irradiation (72 Gy) was given. Although papilledema is a sensitive marker of elevated intracranial pressure, this sign may be masked by constriction of the optic sheath in patients who suffer from leukemic infiltration of the central nervous system and receive high doses of cranial irradiation.

Clinical phenotype of a Japanese family with primary open angle glaucoma caused by a Pro370Leu mutation in the MYOC/TIGR gene.

PURPOSE: To present the phenotype of two patients with primary open angle glaucoma (POAG) caused by a mutation of the myocilin/trabecular meshwork-inducible glucocorticoid response (MYOC/TIGR) gene. METHODS: Complete ocular examinations were performed on the 13-year-old proband, her father, mother, and sister. DNA analysis was performed to detect the mutant gene. RESULTS: The proband and her father were found to have a mutation of the MYOC/TIGR gene. Both patients carried a heterozygous mutation in the 1,109th nucleotide, which corresponds to the 370th amino acid residue of the MYOC/TIGR gene. The clinical characteristics of both patients were: (1) development of POAG at an early age, (2) high peaks of intraocular pressure. and (3) poor response to medical treatment. CONCLUSIONS: The phenotype of these patients with a mutation of the MYOC/TIGR gene agreed with reports of other patients with mutations at other loci in this gene. The discovery of the MYOC/TIGR gene not only makes early detection of glaucoma possible, but also presents a new direction for investigating the pathogenesis of glaucoma.

Menstrual cycle-specific inhibition of the proliferation of endometrial stromal cells by interleukin 6 and its soluble receptor.

OBJECTIVE: This study investigated the possible roles of interleukin 6 and soluble interleukin 6 receptor in the growth of endometrial and endometriotic cells. STUDY DESIGN: Endometrial and endometriotic stromal cells were collected from the uterus or from ovarian chocolate cysts. We examined the effects of interleukin 6, soluble interleukin 6 receptor, and a combination of both factors on the proliferation of endometrial and endometriotic stromal cells. The action of sex steroids on the interleukin 6 regulation of the growth of stromal cells was also evaluated. The gene expressions of interleukin 6 receptor and glycoprotein 130 were examined in endometrial and endometriotic cells by reverse transcription-polymerase chain reaction. RESULTS: Interleukin 6 had no effect on the growth of stromal cells in tissue from the proliferative phase. In contrast, the addition of concentrations of >/=100 pg/mL interleukin 6 induced significant inhibition of stromal cell proliferation in tissue from the secretory phase. Similarly, the addition of soluble interleukin 6 receptor caused significant suppression in the growth of endometrial stromal cells in tissue from the secretory phase but not the proliferative phase. On the other hand, stromal cells of endometriotic tissues were resistant to interleukin 6, showing no inhibitory response. Although the combination treatment did not affect the proliferation of stromal cells of the proliferative phase and of endometriotic tissues, 10 pg/mL interleukin 6 inhibited proliferation of stromal cells of the secretory phase in the presence of 1 ng/mL soluble interleukin 6 receptor. Treatment with estradiol and progesterone for 10 days newly induced the inhibitory response to interleukin 6 in the endometrial cells from the proliferative phase. Expressions of transcripts of interleukin 6 receptor and glycoprotein 130 were observed in the endometrial cells from the proliferative and secretory phases and in endometriotic cells. CONCLUSIONS: Interleukin 6 may play a central role in regulation of the growth of endometrial cells as a mediator of endocrine action. Endometriotic cells may behave differently from their normal counterparts in terms of the inhibitory regulation exerted by interleukin 6.

Role of cytokines in progression of endometriosis.

Peritoneal fluid in women with endometriosis contains an increased number of activated macrophages that secrete a variety of cytokines, including interleukin (IL)-6, IL-8, vascular endothelial growth factor, and tumor necrosis factor-alpha (TNF-alpha). Cytokines may be involved in the control of implantation and the growth of endometrial cells outside the uterus. In addition, several cytokines have been implicated in or directly associated with angiogenic activity in endometriosis. There could be a relationship between the levels of cytokines in the peritoneal fluid of patients with endometriosis and the status of the lesions in such patients. Peritoneal endometriosis can be classified as having red, black, or white lesions. Red lesions are known to be an active form of early endometriosis, because vascularization and mitotic activity are shown to be most prominent in these lesions. We found that the peritoneal fluid levels of TNF-alpha and IL-8 were significantly higher in patients with endometriosis, and correlated with the size and number of active lesions. In addition, TNF-alpha and IL-8 stimulated the growth of ectopic endometrial stromal cells. These cytokines with angiogenic activity may therefore have significant roles in the pathogenesis of endometriosis.

Autocrine effects of transforming growth factor-alpha on the development of preimplantation mouse embryos.

PURPOSE: We wished to explore the role of transforming growth factor (TGF)-alpha in mouse embryonic development. METHODS: We examined the gene expression of TGF-alpha and epidermal growth factor receptor (EGFR) in mouse blastocysts by the reverse transcription-polymerase chain reaction and evaluated the effects of TGF-alpha on the development of preimplantation mouse embryos using TGF-alpha antisense oligodeoxynucleotide. Mouse teratocarcinoma F9 cells were also a subject of this study. RESULTS: Gene transcripts of TGF-alpha and EGFR were present in both blastocysts and F9 cells. TGF-alpha significantly stimulated the rate of blastocoel expansion in early-cavitating blastocysts and the proliferation of F9 cells. Northern blot analysis showed that TGF-alpha gene expression in F9 cells was markedly suppressed in the presence of TGF-alpha antisense oligodeoxynucleotide. TGF-alpha antisense oligonucleotide significantly reduced the rate of blastocoel expansion and the growth of F9 cells. The inhibitory effects of TGF-alpha antisense oligonucleotide on blastocysts and F9 cells were reversed by the addition of TGF-alpha. CONCLUSIONS: The present observations suggest that TGF-alpha acts as an autocrine factor in the development of preimplantation mouse embryos.

Paracrine effects of bFGF and KGF on the process of mouse blastocyst implantation.

Implantation is a complex process that requires the interaction of the blastocyst, and subsequently, that of the developing embryos with the endometrium. Several growth factors and cytokines are involved in implantation, but the details of their actions as related to the regulation of blastocyst implantation remain unclear. In the present study, the RT-PCR method was used to determine the gene expression of basic fibroblast growth factor (bFGF), keratinocyte growth factor (KGF), FGF receptor 1 (FGFR1), FGF receptor 2 (FGFR2), and KGF receptor (KGFR) in mouse embryos and in the stromal and epithelial cells of the uterine endometrium. Basic FGF and KGF mRNA were expressed in the endometrial cells, but were not expressed in the embryos. The mRNAs of receptors for bFGF and KGF were expressed in the blastocysts and in the in vitro implanting embryos, suggesting that bFGF and KGF may exert paracrine effects on blastocyst implantation. In this mouse model of blastocyst implantation, it was found that transforming growth factor alpha (TGF-alpha) at the concentrations of 1 ng/ml and 10 ng/ml significantly enhanced the blastocyst attachment and trophoblast spreading and increased trophoblast surface area. Relatively high concentrations of bFGF (100-500 ng/ml) significantly enhanced the rates of blastocyst attachment and of trophoblast spreading and promoted the expansion of the surface area of the implanting embryos. Unlike the rates of blastocyst attachment and trophoblast spreading, the surface area of the spreading embryos was significantly increased by addition of KGF (1-100 ng/ml). These results suggest that the bFGF and KGF derived from the endometrial cells exert paracrine effects on the process of implantation by stimulating trophoblast outgrowth through their cognate receptors.

Systemic administration of rIL-12 synergistically enhances the therapeutic effect of a TNF gene-transduced cancer vaccine.

Interleukin-12 (IL-12) is a potent antitumor cytokine, which induces and enhances the activity of natural killer (NK) cells, lymphokine activated killer (LAK) cells and cytotoxic T lymphocytes (CTL). IL-12 also stimulates IFN-gamma production from both T cells and NK cells. In this study, we transfected methylcholanthrene-induced fibrosarcoma (MCA-D) with TNF gene and investigated the therapeutic effect of TNF gene-transduced cancer vaccine and whether the vaccination effect is enhanced by systemic administration of recombinant IL-12 (rIL-12), in a murine model. TNF gene-transduced cancer vaccine or systemic administration of rIL-12 showed slight or moderate inhibition of pre-established tumor. However, simultaneous application of the vaccine and rIL-12 resulted in complete eradication. The cytotoxicity of CTL against parental tumor cells was enhanced with the combination of the vaccine and rIL-12, and IFN-gamma production from spleen cells also increased synergistically. Our findings show that synergistic enhancement of CTL activity and IFN-gamma production could play an important role in the antitumor effect of combination therapy using TNF gene-transduced cancer vaccine and rIL-12.