Preoperative diagnosis of colouterine fistula secondary to diverticulitis by sonohysterography with contrast medium.

Colouterine fistulae secondary to sigmoid diverticulitis are unusual. Methods for diagnosis remain to be established. We report a case with a colouterine fistula in which sonohysterography detected the flow of ultrasound contrast medium between the uterine cavity and the sigmoid colon through the posterior uterine wall, thus confirming the diagnosis. The diagnosis was further substantiated by a charcoal challenge test. The patient underwent en bloc resection of the uterus, Fallopian tubes, ovaries and sigmoid colon, the organs involved with diverticulitis. This is the first report to describe a colouterine fistula successfully diagnosed by sonohysterography using ultrasound contrast medium.

Inhibition of nitric oxide synthesis potentiates apoptosis in the rabbit corpus luteum.

To determine if nitric oxide (NO) plays a role in corpus luteum (CL) physiology by affecting progesterone secretion or luteal apoptosis, an in-vitro pseudopregnant rabbit ovarian perfusion system was used to measure the effects of an inhibitor of NO synthesis, NG-nitro-L-arginine methyl ester (L-NAME), on progesterone secretion and corpus luteal apoptosis as measured by internucleosomal DNA breakdown. Pseudopregnant rabbit ovaries perfused in vitro with L-NAME did not demonstrate any significant differences compared with control ovaries in progesterone secretion. However, apoptosis, as measured by internucleosomal breakdown, was significantly increased in L-NAME-perfused CL compared with controls. While NO does not appear to directly affect progesterone secretion, there does appear to be a role for NO in CL maintenance, or a role for inhibition of NO production in CL regression.

Abnormal Fhit expression is an independent poor prognostic factor for cervical cancer.

We analysed the expression of the fragile histidine triad (FHIT) gene in cervical cancer to evaluate its clinical relevance in relation to human papillomavirus (HPV) infection. A total of 73 women with cervical cancer of stage Ib or more advanced (67 squamous cell carcionomas, four adenocarcinomas, two adenosquamous carcinomas) were examined for Fhit expression by immunohistochemistry. They were further analysed for the presence of HPV and its subtype. Abnormal expression of Fhit (absent or reduced Fhit expression) was observed in 52 cases (71.2%). The high-risk HPV DNAs for cervical cancer, including type 16, 18, 31, 33, 51, 52, 58, 68, were identified in 63 cases (86%). The abnormal Fhit expression was not related to the clinicopathological factors including histology, tumour stage, and HPV type. Notably, the 5-year survival of patients showing the abnormal Fhit expression was significantly poorer than those showing normal Fhit expression (64 versus 87%, P=0.035). Interestingly, the mean age of the patients with the abnormal Fhit expression was significantly less than those with the normal Fhit expression (51.6 versus 58.7 years of age, P=0.027, student's t-test). These data imply that the aberrant Fhit expression could be a poor prognostic factor independent of HPV. In the light of a high incidence of abnormal Fhit expression in younger patients and HPV as a key player in cervical carcinogenesis, abnormal Fhit expression may accelerate carcinogenesis in concert with HPV.

Effects of gonadotropin-releasing hormone analog treatment on skin condition.

The skin is a target organ of estrogens. Thus, theoretically, a hypoestrogenic state induced by gonadotropin-releasing hormone analog (GnRHa) treatment may have effects on skin condition. The aim of this study was to evaluate skin condition during GnRHa treatment. Sixteen premenopausal women undergoing GnRHa treatment for 16 weeks, as a presurgical treatment for uterine leiomyomas, were studied. Measurement of serum estradiol levels and epidermal hydration, and evaluation of subjective findings on skin condition using a questionnaire, were performed every 4 weeks during the treatment period. Serum estradiol levels were significantly suppressed at 4 weeks of treatment, and remained low afterwards. Epidermal hydration measured by corneometer did not show any significant difference at any time point examined, compared with that before treatment. No particular subjective findings relating to the skin (dryness, wrinkling, roughness, pigmentation, itching, formication, reaction to cosmetics) were reported during treatment, whereas complaints about hot flushes and sweating were notable. The results of this preliminary study support the notion that GnRHa treatment for 16 weeks is unassociated with apparent changes in skin condition.

Demonstration of angiogenin in human endometrium and its enhanced expression in endometrial tissues in the secretory phase and the decidua.

Angiogenesis is thought to be crucial for normal physiology of the endometrium, where dynamic vascular remodeling occurs during the menstrual cycle and pregnancy. We investigated the presence of angiogenin, a potent inducer of angiogenesis, and the regulatory mechanisms of its production in the human endometrium. Western blot analysis demonstrated that angiogenin protein expression increased by 3- to 4-fold in the endometrium in the mid and late secretory phases and in early gestation relative to that during the proliferative phase. Quantitative mRNA analysis showed the similar tendency in the expression of angiogenin mRNA in the endometrium, with the highest levels observed in the mid and late secretory phases and early gestation. An immunohistochemical study showed that angiogenin was expressed in both stromal cells and epithelial cells, with indistinguishable intensity between these cells regardless of phases of the menstrual cycle. In support of the Western blot analysis, the intensity of staining appeared to be highest in the mid to late secretory phases relative to other phases. Consistent with these in vivo results, decidualized cultured stromal cells, after treatment with progesterone or progesterone plus E2, exhibited the capacity to secrete significantly increased amounts of angiogenin compared with untreated or E2 alone-treated control group. Both the treatment with (Bu)2cAMP and hypoxic conditions stimulated angiogenin secretion by stromal cells. For isolated epithelial cells, hypoxia stimulated angiogenin secretion, whereas (Bu)2cAMP had no appreciable effect. In summary, we demonstrated the presence of angiogenin in human endometrium and its possible local regulatory factors, such as progesterone, cAMP, and hypoxia. These findings along with its enhanced expression in the endometrium in the secretory phase and in decidual tissues raise the possibility that angiogenin may play a role in establishing pregnancy.

Usefulness of long-jaw forceps in laparoscopic cornual resection of interstitial pregnancies.

Although laparoscopic surgery could be an option for treating interstitial pregnancies, it has not gained wide acceptance largely because of difficulty achieving hemostasis. To overcome this, we employed long-jaw forceps in three cases of interstitial pregnancy that were successfully treated by laparoscopic cornual resection. The forceps grasped a relatively large portion of pregnant myometrium without slipping, thus securing hemostasis and facilitating suturing.

Evidence for the presence of keratinocyte growth factor (KGF) in human ovarian follicles.

The presence of keratinocyte growth factor (KGF) in human follicular fluid (FF) was investigated in a total of 145 FFs obtained during oocyte retrieval for in vitro fertilization (IVF) from 29 patients with no apparent endocrine disorders. The concentrations of KGF, estradiol, progesterone, testosterone and human chorionic gonadotropin (hCG) in FF were measured by enzyme-linked immunosorbent assay. FF samples contained relatively higher amounts of KGF (2194+/-87 pg/ml), whereas its concentrations in serum were below assay limit (<31.2 pg/ml). Concentrations of KGF in FF were positively correlated with both progesterone (r=0.311, p<0.0005) and testosterone (r=0.230, p<0.01) concentrations in FF. However, KGF concentrations were not significantly correlated with estradiol and hCG concentrations. KGF in FF was detected as a broad band (26-29 kD) by immunoblotting, the size being reduced by 7kD after N-glycosidase treatment. In an in vitro experiment, KGF suppressed the basal and hCG-stimulated progesterone production by cultured human luteinized granulosa cells. summary, we demonstrated the presence of KGF in human ovarian follicles, suggesting its possible role as a local factor in regulating human ovarian functions.

Evidence for the expression of interleukin (IL)-18, IL-18 receptor and IL-18 binding protein in the human endometrium.

To see whether the interleukin (IL)-18 system is operative in the endometrium, we examined the expression of IL-18, IL-18 receptor (IL-18R) and IL-18 binding protein (IL-18BP), the substance known to neutralize IL-18 activity, in this tissue. Reverse transcription-polymerase chain reaction analyses showed that IL-18, IL-18R and IL-18BP mRNA were constitutively expressed without significant fluctuation throughout the menstrual cycle. When epithelial cells and stromal cells were cultured separately, the expression levels of IL-18 mRNA in epithelial cells were about 18-fold higher compared to those in stromal cells. Furthermore, the IL-18 precursor protein was detected by Western blot analysis in cultured epithelial cells but not in stromal cells. Recombinant human IL-18 stimulated the secretion of interferon (IFN)-gamma by resident bone marrow-derived cells in the endometrium. On the other hand, IFN-gamma up-regulated the IL-18BP expression both in cultured epithelial cells and stromal cells. Thus, we have presented evidence for the presence of the IL-18 system in the human endometrium. In light of its immunomodulatory roles in a variety of tissues, this system may afford protection against pathogenic micro-organisms and provide a regulatory mechanism for controlled trophoblast invasion by modulating a local cytokine network.

Hyperemesis gravidarum associated with thyrotoxicosis and a past history of an eating disorder.

We present a case of severe hyperemesis gravidarum (HG) associated with thyrotoxicosis in a woman with a past history of an eating disorder. She had developed persistent HG from early pregnancy until about at the end of the second trimester with a body loss of 14 kg. Total parenteral nutrition was effective in alleviateing HG. It is suggested that even a past history of an eating disorder could be at risk of developing HG.

Is an elevation in basal follicle-stimulating hormone levels in unexplained infertility predictive of fecundity regardless of age?

An elevation in follicle-stimulating hormone (FSH) levels is considered to reflect lowered ovarian function, resulting in poor fecundity in infertile women. However, it remains to be clarified whether or not the significance of FSH levels applies equally to all women irrespective of age. The objective of the present study is to compare basal FSH levels in infertile women who conceived or not after stratification by age. A total of 144 infertile women between ages 25 and 45 who underwent infertility treatment due to unexplained infertility in the University of Tokyo Hospital were included in the retrospective study. Subjects were divided by age into two groups, < 38 (n=98) vs > or = 38 (n=46) years, with ages ranging from 25 to 37, and from 38 to 45, respectively. Blood samples were collected in early follicular phase (day 4-6) for assessment of basal levels of LH, FSH, and PRL. In the older group, pregnant cases had significantly lower FSH levels (6.07 +/- 2.83 mIU/ml) than nonpregnant cases (9.60 +/- 3.67 mIU/ml), whereas no difference in basal FSH levels was observed between pregnant and nonpregnant cases in the younger group. Basal FSH levels of pregnant cases in the older group were significantly lower than those of pregnant cases in the younger group (8.26 +/- 2.95 mIU/ml). Basal LH and PRL levels were not related to fecundity in either group. Thus, an increase in basal FSH levels as a predictor of fecundity should be considered in the context of age.

Inhibition of human epithelial ovarian cancer cell growth in vitro by somatostatin analog RC-160.

In this study, we investigated the effects of somatostatin analog RC-160 on the growth of the OV-1063 human epithelial ovarian cancer cell line in vitro. RC-160 inhibited cell proliferation, as measured by cell number, and [(3)H]thymidine incorporation into DNA at 10(-9)-10(-5) M. In OV-1063 cells, (125)I-labeled RC-160 was bound to one class of specific, saturable binding sites with high affinity (K(d) = 0.2 +/- 0.03 nM) and low capacity (5,500 binding sites per cell). (125)I-labeled RC-160 could be displaced by unlabeled RC-160. Ligand binding was dependent on time and temperature. Receptor internalization assay showed that the ligand-receptor complex was internalized at 37 degrees C, which indicates the presence of biologically active somatostatin receptors on OV-1063 cells. These results suggest that somatostatin analog RC-160 can suppress the growth of OV-1063 human epithelial ovarian cancer cells by a direct action and that the inhibitory effect of somatostatin analog is mediated through the high-affinity somatostatin receptors.

Increased concentrations of soluble tumour necrosis factor receptor (sTNFR) I and II in peritoneal fluid from women with endometriosis.

Tumour necrosis factor alpha (TNFalpha), a proapoptotic cytokine, is known to be present in peritoneal fluid from women with endometriosis. An emerging view is that soluble TNF receptors (sTNFR) can modulate the effects of TNFalpha by acting as TNFalpha antagonists. To assess the relevance of sTNFRs in the pathophysiology of endometriosis, concentrations of sTNFR I, sTNFR II and TNFalpha in peritoneal fluid from women with endometriosis (n = 53) and without endometriosis (n = 40) were measured. Concentrations of both sTNFR I and sTNFR II in peritoneal fluid from women with endometriosis were significantly higher than in peritoneal fluid from women without endometriosis, both in the follicular and the luteal phases. TNFalpha concentrations did not differ in patients with and without endometriosis in both phases. When stratified by the stage of the disease, women with both stages I/II and stages III/IV exhibited significantly higher concentrations of sTNFR I and sTNFR II in peritoneal fluid, compared with women without endometriosis, whereas no appreciable difference in the concentrations was detected between stages I/II and stages III/IV. A significant correlation was found between the concentrations of sTNFR I and sTNFR II; while the correlations between TNFalpha and sTNFR I or sTNFR II, were either not significant or were very weak. Furthermore, mRNA for the membrane-associated TNF receptor type 1 and TNF receptor type 2, both of which convey the effects of TNFalpha, were shown to be expressed in endometriotic tissues as well as eutopic endometrium. Together, these findings suggest a possible involvement of sTNFRs in the pathophysiology of endometriosis.

Evidence for the presence of angiogenin in human follicular fluid and the up-regulation of its production by human chorionic gonadotropin and hypoxia.

Angiogenesis is an essential event during the development of the ovarian follicle and ensuing formation of the corpus luteum. We investigated the presence of angiogenin, a potent inducer of angiogenesis, and the regulatory mechanisms of its production in the human ovary. Follicular fluid (FF) and granulosa cells (GCs) were collected from women undergoing in vitro fertilization and embryo transfer. The presence of angiogenin in FF and GCs was demonstrated by Western blot analysis. The production of angiogenin by cultured GCs was stimulated with the addition of human CG or cAMP or under the hypoxic milieu. Concentrations of angiogenin in FF from an individual follicle were positively correlated with those of progesterone, but not estradiol and testosterone. Given the presence of angiogenin in FF and up-regulation of its production by human CG and hypoxia, it seems logical to assume that angiogenin may play a role as a local angiogenic factor in the human ovary.

Regulation of nitric oxide synthase to promote cytostasis in ovarian follicular development.

Our own recent studies have demonstrated that inducible nitric oxide synthase (iNOS) is predominantly localized in granulosa cells of healthy immature follicles in the rat ovary, whereas granulosa cells of either healthy mature follicles or follicles destined to be atretic are devoid of iNOS. These findings suggest that iNOS is pivotal for immature follicles to remain dormant. To test this hypothesis, we examined the effects of a GnRH agonist (buserelin), a proapoptotic substance, and epidermal growth factor (EGF), a mitogenic and, consequently, antiapoptotic factor, on the amount of iNOS mRNA in rat granulosa cells. Administration of buserelin in immature female rats transiently diminished iNOS mRNA levels in the ovaries as determined by Northern blot analysis. In cultured rat granulosa cells, buserelin and EGF increased the incidence of apoptosis and DNA synthesis, respectively, whereas both reduced iNOS mRNA levels as determined by reverse transcription-coupled polymerase chain reaction. The concomitant addition of S-nitroso-N-acetyl-DL-penicillamine, an NO donor, together with buserelin or EGF eliminated the observed effects of these substances (i.e., induction of apoptosis and stimulation of DNA synthesis, respectively). These results suggest that the changes in developmental status of immature follicles either into development or atresia are associated with reduced iNOS levels in granulosa cells, thus reinforcing the notion of NO as a cytostatic factor in ovarian follicles.

Estradiol-mediated suppression of apoptosis in the rabbit corpus luteum is associated with a shift in expression of bcl-2 family members favoring cellular survival.

In the rabbit, estradiol is the primary luteotropic hormone. Estradiol withdrawal results in a rapid decline in serum progesterone and eventually in corpus luteum (CL) regression. The objective of this study was to determine whether estradiol modulates luteal cell apoptosis. In the first experiment, rabbits were randomly assigned to one of five experimental groups. An empty capsule (control) or estradiol-filled Silastic capsule was inserted s.c. on Day 0 of pseudopregnancy (day of hCG administration). On Day 11 of pseudopregnancy, some of the group I (control) and group II (estradiol capsule) rabbits were subjected to laparotomy, and one ovary from each rabbit was perfused in vitro to determine progesterone secretion rates. The CL from the contralateral ovary were dissected, snap-frozen, and stored at -70 degrees C until analyzed for internucleosomal DNA cleavage (apoptosis). Estradiol-containing capsules were removed from some of the remaining rabbits on Days 8, 9, and 10 to initiate estradiol deprivation. Rabbits were then subjected to laparotomy 24, 48, or 72 h after capsule removal (groups III, IV, and V, respectively), and ovaries or CL were processed as described above. Deprivation of estradiol for 24 (group III), 48 (group IV), or 72 (group V) h in vivo reduced in vitro progesterone secretion rates by more than 90% as compared to that in ovaries collected from estradiol capsule-intact animals. After in vivo endogenous estradiol suppression, withdrawal of exogenous estradiol resulted in luteal cell apoptosis, which increased in a time-dependent manner. Northern blot analysis revealed an increase in bax mRNA levels and a decrease in bcl-x mRNA levels coincident with luteal cell apoptosis induced by estradiol withdrawal. These data demonstrate that changes in progesterone production caused by estradiol exposure and deprivation are in part related to luteal cell apoptosis, and alterations in the expression of bcl-2 gene family members may be one of the mechanisms by which estradiol exerts its luteotropic effect in the rabbit CL.

High and low molecular weight DNA cleavage in ovarian granulosa cells: characterization and protease modulation in intact cells and in cell-free nuclear autodigestion assays.

To continue elucidation of the biochemical and molecular pathways involved in the induction of apoptosis in granulosa cells (GC) of ovarian follicles destined for atresia, we characterized the occurrence and protease modulation of high and low molecular weight (MW) DNA fragmentation during rat GC death. Atresia of ovarian follicles, occurring either spontaneously in vivo or induced in vitro, was associated with both high MW and internucleosomal (low MW) DNA cleavage. Incubation of follicles in the presence of a putative irreversible and non-competitive inhibitor of caspase-1 (interleukin-1beta-converting enzyme or ICE), sodium aurothiomalate (SAM), completely prevented internucleosomal, but not high MW, DNA cleavage. As reported previously, morphological features of apoptosis (pyknosis, cellular condensation) and atresia (granulosa cell disorganization, oocyte pseudomaturation) remained detectable in SAM-treated follicles. The potential involvement of proteases in endonuclease activation was further analyzed in cell-free assays using nuclei from both GC (which autodigest their DNA) and HeLa cells (HC, which do not autodigest their DNA unless incubated with extracts prepared from other cell types). Crude cytoplasmic extracts prepared from GC induced both high MW and internucleosomal DNA cleavage in HC nuclei. The induction of low, but not high, MW DNA cleavage in HC nuclei by GC extracts was suppressed by pretreatment of the extracts with SAM or with any one of the serine protease inhibitors, dichloroisocoumarin (DCI), N-tosyl-L-leucylchloromethylketone (TLCK) or N-tosyl-L-phenylchloromethylketone (TPCK). Interestingly, SAM and DCI also prevented cation-induced low MW DNA fragmentation in GC nuclei; however, TLCK and TPCK were without effect. Our results support a role for cytoplasmic and nuclear serine proteases in the activation of the endonuclease(s) responsible for internucleosomal DNA cleavage during apoptosis.

Analysis of apoptosis and expression of bcl-2 gene family members in the human and baboon ovary.

Recent data support a role for apoptosis, under tight regulatory control by bcl-2, oxidative stress response, tumor suppressor, and CASP gene family members, in mediating granulosa cell demise during follicular atresia in the rodent and avian ovary. Herein we evaluated the occurrence of apoptosis in the human and baboon ovary relative to follicular health status, and analyzed expression of several cell death genes in these tissues. In situlocalization of DNA strand breaks in fixed human and baboon ovarian tissue sections indicated that apoptosis was essentially restricted to granulosa cells of atretic antral follicles. Biochemical analysis of DNA oligonucleosomes in individual follicles isolated from baboon ovaries during the ovulatory phase revealed the presence of apoptotic DNA fragments in subordinate but not dominant follicles, thus substantiating the in situ labeling studies. Messenger RNA transcripts encoded by the bax death susceptibility gene, the bcl-xlong survival gene, the bcl-xshort pro-apoptosis gene, the p53 tumor suppressor gene, and two members of the CASP gene family (CASP-2/Ich-1, CASP-3/CPP32), were detected by Northern blot analysis of total RNA prepared either from human ovaries or from Percoll-purified granulosa-lutein cells obtained from patients undergoing assisted reproductive technologies. Lastly, immunohistochemical localization of the BAX death-susceptibility protein in the human ovary revealed abundant expression in granulosa cells of early atretic follicles, whereas BAX protein was extremely low or non-detectable in healthy or grossly-atretic follicles. We conclude that apoptosis occurs during, and is probably responsible for, folicular atresia in the human and baboon ovary. Moreover, apoptosis in the human ovary is likely controlled by altered expression of the same cohort of cell death regulatory factors recently implicated as primary determinants of apoptosis induction or suppression in the rodent ovary.

Interleukin-1 beta-converting enzyme-related proteases (IRPs) and mammalian cell death: dissociation of IRP-induced oligonucleosomal endonuclease activity from morphological apoptosis in granulosa cells of the ovarian follicle.

The Caenorhabditis elegans death susceptibility gene, ced-3, has a number of homologs in vertebrate species, including interleukin-1 beta (IL-1 beta)-converting enzyme (ICE), Ich-1long, and CPP32. These genes, which encode a family of related proteases, have been shown to induce apoptosis when transfected into eukaryotic cells. However, it remains to be determined whether these proteases are involved in apoptotic cell death under physiological conditions. The purpose of these studies was to examine the role of ICE-related proteases (IRPs) in apoptosis using a physiologically relevant model system, the ovarian follicle. Somatic granulosa cells within ovarian follicles undergo apoptosis during follicular atresia, a process responsible for the depletion of greater than 95% of the follicles established in the postnatal ovary. To accomplish these studies, we cloned partial rat complementary DNAs encoding ICE, Ich-1, and CPP32 and used these complementary DNAs to examine the gonadotropin regulation of ICE, Ich-1, and CPP32 gene expression in the immature rat ovary. We also examined levels of ICE activity in healthy and atretic rat follicles by monitoring the conversion of exogenous pro-IL-1 beta to the active cytokine, and then evaluated the actions of recombinant IL-1 beta on apoptosis in follicles incubated in vitro. Finally, we tested the requirement for IRP activity in granulosa cell apoptosis and follicular atresia by incubating follicles without and with IRP inhibitors. Northern blot analysis of total RNA samples indicated that gonadotropin-promoted follicular survival was associated with reduced ovarian expression of messenger RNAs encoding Ich-1 and CPP32. In contrast, ICE messenger RNA levels were extremely low and were not affected by gonadotropin treatment. We were also unable to detect ICE activity in proteins extracted from either healthy or atretic rat follicles, collectively suggesting that ICE per se may not function in granulosa cell death. As another approach to determine whether ICE is involved in atresia, healthy antral follicles were isolated from ovaries of gonadotropin-primed immature rats and incubated for 24 h in the absence or presence of 100 ng/ml transforming growth factor-alpha (TGF alpha) without and with 100 ng/ml IL-1 beta. Granulosa cells within follicles incubated in medium alone exhibited extensive levels of apoptosis, and this onset of apoptosis was prevented by the inclusion of TGF alpha. Addition of IL-1 beta did not alter basal levels of apoptosis nor did the cytokine antagonize TGF-alpha-promoted follicle survival, providing additional evidence that ICE activity is not required for atresia to occur.(ABSTRACT TRUNCATED AT 400 WORDS)

Vasoactive intestinal peptide-mediated suppression of apoptosis in the ovary: potential mechanisms of action and evidence of a conserved antiatretogenic role through evolution.

Vasoactive intestinal peptide (VIP)-containing nerve fibers are present in ovarian follicles at all stages of development, and VIP, acting primarily via the cAMP pathway, has been reported to modulate many aspects of granulosa cell function. Herein we examined the effects of VIP and its potential mechanisms of action on apoptosis in antral follicles isolated from ovaries of gonadotropin-primed immature rats and incubated in vitro under serum-free conditions. Additionally, the effects of VIP on apoptosis in isolated avian granulosa cells incubated in vitro were used as a comparative model system to determine whether the ability of VIP to modulate apoptosis in the ovary has been conserved through evolution. Genomic DNA extracted from incubated rat antral follicles exhibited extensive levels of internucleosomal DNA cleavage characteristic of cell death via apoptosis. Treatment of follicles with VIP (1-1000 nM) caused a dose-dependent reduction in the extent of apoptotic DNA breakdown, with a maximal effect achieved with 100 nM VIP. Provision of the adenylyl cyclase activator, forskolin (10 microM), mimicked the inhibitory effect of VIP on apoptosis and concomitantly increased intrafollicular cAMP accumulation, suggesting a role for the cAMP pathway in mediating the immediate actions of VIP on follicular cell survival. Moreover, treatment of rat antral follicles with insulin-like growth factor-binding protein 3 (3 micrograms/ml) partially antagonized the ability of VIP (100 nM) to suppress apoptosis, suggesting involvement of endogenous insulin-like growth factor I in mediating the downstream actions of VIP in incubated rat antral follicles. To further confirm that VIP and activation of the cAMP pathway prevented atresia, individual rat antral follicles incubated for 24 h in the absence or presence of VIP (100 nM) or forskolin (10 microM) were fixed, embedded, and sectioned for morphological analysis. Follicles fixed immediately after isolation from equine CG-primed rat ovaries were classified as morphologically healthy, consistent with the absence of biochemical evidence for apoptosis (e.g. oligonucleosomes) in this pool of follicles. Follicles incubated for 24 h in the absence of tropic support displayed extensive granulosa cell pyknosis and disorganization characteristic of follicles at a moderate stage of atresia. Inclusion of VIP or forskolin maintained the morphological health status of incubated follicles at that resembling healthy follicles fixed immediately after isolation from ovaries of equine CG-primed rats. Lastly, extensive levels of internucleosomal DNA cleavage were also detected in avian granulosa cells incubated for 6 h under serum-free conditions.(ABSTRACT TRUNCATED AT 400 WORDS)

Tentative diet for liver failure containing well-polished rice.

A liver-failure diet (low in protein) that contained rice polished to 50% to reduce the protein content of the diet was given to patients with uncompensated liver cirrhosis and compared with a standard liver-failure diet containing conventionally processed rice. The amount of boiled rice served in each meal could be increased by using well-polished rice and the use of supplementary sources of energy (powdered starch syrup, jelly, cookies, and candy sugar) was unnecessary. In the liver-failure diet containing well-polished rice, the methionine contents could be reduced and the Fischer ratio could be increased. The ingestion rate of the diet with well-polished rice was 80% and the diet was rated favorably in a questionnaire on palatability. Decreases in blood ammonia concentrations were observed in three patients given the liver-failure diet with well-polished rice for 2 wk by the crossover method.