Multivariate curve resolution using a combination of mid-infrared and near-infrared spectra for the analysis of isothermal epoxy curing reaction.

Multivariate curve resolution (MCR) was applied to a hetero-spectrally combined dataset consisting of mid-infrared (MIR) and near-infrared (NIR) spectra collected during the isothermal curing reaction of an epoxy resin. An epoxy monomer, bisphenol A diglycidyl ether (BADGE), and a hardening agent, 4,4'-diaminodiphenyl methane (DDM), were used for the reaction. The fundamental modes of the NH and OH stretches were highly overlapped in the MIR region, while their first overtones could be independently identified in the NIR region. The concentration profiles obtained by MCR using the hetero-spectral combination showed good agreement with the results of calculations based on the Beer-Lambert law and the mass balance. The band assignments and absorption sites estimated by the analysis also showed good agreement with the results using two-dimensional (2D) hetero-correlation spectroscopy.

Expression of steroidogenic factor 1 and pituitary specific transcription factor 1 in rat pituitary adenomas.

The protein expressions of steroidogenic factor l (SF-l) and pituitary-specific transcription factor 1 (Pit-1) were investigated immunohistochemically for 53 spontaneous pituitary adenomas of the pars distalis from male Crl:CD(SD) rats. Luteinizing hormone (LH)-positive/prolactin (PRL)-negative and LH-negative/PRL-positive adenomas showed that the expression of SF-1 and Pit-1 was exclusively related to the immunoreactivity of LH and PRL, respectively. All double-positive adenomas (positive for both LH and PRL) were positive for Pit-1 and were supposed to be derived from PRL cells, although some of them also showed SF-1 immunoreactivity. In addition, all null cell adenomas (negative for all anterior pituitary hormones) were positive for SF-1 and negative for Pit-1, indicating that they originated from the gonadotroph cell lineage. This is the first report focusing on the application of transcription factors for the classification of rat pituitary adenomas.

Antitumor activity and pharmacodynamic biomarkers of a novel and orally available small-molecule antagonist of inhibitor of apoptosis proteins.

Inhibitor of apoptosis proteins (IAP), which are key regulators of apoptosis, are inhibited by second mitochondria-derived activator of caspase (SMAC). Small-molecule IAP antagonists have recently been reported as novel therapeutic treatments for cancer. In this study, we showed that the octahydro-pyrrolo[1,2-a]pyrazine derivative, T-3256336, is a novel and orally available small-molecule IAP antagonist. T-3256336 selectively binds to and antagonizes protein interactions involving cellular IAP-1 (cIAP-1), cIAP-2, and X-linked IAP (XIAP). T-3256336 induced the rapid proteasomal degradation of cIAP-1 and activated TNF-α-dependent extrinsic apoptosis signaling in cultured cells. In a MDA-MB-231-Luc breast cancer xenograft model, T-3256336 induced cIAP-1 degradation, TNF-α production, and caspase activation in tumors, which resulted in strong antitumor activities. T-3256336 induced increases in the plasma levels of TNF-α and fragmented cytokeratin-18, which correlated with the antitumor potency in MDA-MB-231-Luc xenograft models. This study provided further insights into biomarkers of IAP antagonists. Furthermore, our data provided evidence that T-3256336 is a promising new anticancer drug worthy of further evaluation and development.

Two-step curing reaction of epoxy resin studied by thermal analysis and infrared spectroscopy.

A curing reaction of bisphenol A diglycidyl ether epoxy resin with 4,4'-diaminodicyclohexyl methane hardener was investigated by means of modulated differential scanning calorimetry (MDSC) and infrared (IR) spectroscopy. MDSC observation revealed that the curing process of the resin occurred in two steps. Mid-infrared and near-infrared spectra of the resin were measured as a function of temperature. The obtained spectra were analyzed by perturbation-correlation moving-window two-dimensional correlation spectroscopy (PCMW2D-COS). The first step was revealed as a polymerization reaction among the oxirane group and primary and secondary amine groups, followed by etherification; the second step of the curing process occurred in the vicinity of the gelation point and was characterized by the growth of a three-dimensional cross-linking structure with tertiary amine and etherification of the hydroxyl group.

Large parasitic myomas in abdominal subcutaneous adipose tissue along a previous myomectomy scar.

The incidence of iatrogenic parasitic uterine myomas associated with the use of a laparoscopic morcellator has been increasing over the past decade. Recently, we encountered a very rare case with a large parasitic myoma measuring 12 cm in diameter in the abdominal subcutaneous adipose tissue along an abdominal longitudinal surgical scar. The patient had twice undergone abdominal myomectomy for multiple fundal myomas. This is the first report describing a case with as large a parasitic myoma presenting in the suprafascial adipose tissue under the surgical scar after laparotomy. In such a case demonstrating a solid tumor of unknown cause after a gynecologic surgical procedure, a parasitic myoma must be included in the differential diagnosis.

Induction and persistence of abnormal testicular germ cells following gestational exposure to di-(n-butyl) phthalate in p53-null mice.

Phthalate esters are commonly used plasticizers found in many household items, personal care products, and medical devices. Animal studies have shown that in utero exposure to di-(n-butyl) phthalate (DBP) within a critical window during gestation causes male reproductive tract abnormalities resembling testicular dysgenesis syndrome. Our studies utilized p53-deficient mice for their ability to display greater resistance to apoptosis during development. This model was chosen to determine whether multinucleated germ cells (MNG) induced by gestational DBP exposure could survive postnatally and evolve into testicular germ cell cancer. Pregnant dams were exposed to DBP (500 mg/kg/day) by oral gavage from gestational day 12 until birth. Perinatal effects were assessed on gestational day 19 and postnatal days 1, 4, 7, and 10 for the number of MNGs present in control and DBP-treated p53-heterozygous and null animals. As expected, DBP exposure induced MNGs, with greater numbers found in p53-null mice. Additionally, there was a time-dependent decrease in the incidence of MNGs during the early postnatal period. Histologic examination of adult mice exposed in utero to DBP revealed persistence of abnormal germ cells only in DBP-treated p53-null mice, not in p53-heterozygous or wild-type mice. Immunohistochemical staining of perinatal MNGs and adult abnormal germ cells was negative for both octamer-binding protein 3/4 and placental alkaline phosphatase. This unique model identified a role for p53 in the perinatal apoptosis of DBP-induced MNGs and provided insight into the long-term effects of gestational DBP exposure within a p53-null environment.

Fourteen-year-old girl with recurrent luteal-phase-dependent episodes of high fever.

Recurrent episodes of high fever during the luteal phase are exceedingly rare. A 14-year-old girl with a 1-year history of recurrent febrile episodes associated with the menstrual cycle was referred to our department. Febrile episode (39-41°C) occurred for 10 to 12 days concomitantly with the luteal phase of each ovulatory cycle. To suppress the ovulatory cycle, gonadotropin releasing hormone agonist was administered for 5 months, and then febrile attack disappeared (34.5-36.2°C). Before and during gonadotropin releasing hormone agonist therapy, serum levels of inflammatory cytokines (interleukin [IL]-1ß, IL-2, IL-6, IL-8, IL-10, and tumor necrosis factor-α) were unchanged and all of these were within the normal ranges. In our case, luteal-phase-dependent febrile episodes may not be attributable to the intervention of inflammatory cytokines as major fever mediators.

Salivary mucocele in a laboratory beagle.

The histologic characteristics of a salivary mucocele in a beagle used in a toxicity study are described in this report. A pale yellowish cyst under the mandibular skin containing frothy mucus was observed at necropsy. Microscopically, numerous villous projections arose from the internal surface of the cyst and were lined by stratified epithelial-like macrophages, which were immunopositive for macrophage scavenger receptor A. A ruptured sublingual interlobar duct connected to the lumen was observed near the cyst. Luminal amorphous material showed a positive reaction with Alcian blue and periodic acid-Schiff staining as did mucin in the sublingual gland. Ultrastructurally, the epithelial-like macrophages had numerous vacuoles containing electron-lucent material, which was presumed to be lysosomal in origin, and had pseudopods on their cell surfaces interdigitating with those on the adjacent cells. This case report helps to understand the diversity of the background findings in beagles used in toxicity studies.

Suppression of radiation-induced testicular germ cell apoptosis by 2,5-hexanedione pretreatment. III. Candidate gene analysis identifies a role for fas in the attenuation of X-ray-induced apoptosis.

Germ cell apoptosis directly induced by x-radiation (x-ray) exposure is stage specific, with a higher incidence in stage II/III seminiferous tubules. A priming exposure to the Sertoli cell toxicant 2,5-hexanedione (HD) results in a marked reduction in x-ray-induced germ cell apoptosis in these affected stages. Because of the stage specificity of these responses, examination of associated gene expression in whole testis tissue has clear limitations. Laser capture microdissection (LCM) of specific cell populations in the testis is a valuable technique for investigating the responses of different cell types following toxicant exposure. LCM coupled with quantitative real-time PCR was performed to examine the expression of apoptosis-related genes at both early (3 h) and later (12 h) time points after x-ray exposure, with or without the priming exposure to HD. The mRNAs examined include Fas, FasL, caspase 3, bcl-2, p53, PUMA, and AEN, which were identified either by literature searches or microarray analysis. Group 1 seminiferous tubules (stages I-VI) exhibited the greatest changes in gene expression. Further analysis of this stage group (SG) revealed that Fas induction by x-ray is significantly attenuated by HD co-exposure. Selecting only for germ cells from seminiferous tubules of the most sensitive SG has provided further insight into the mechanisms involved in the co-exposure response. It is hypothesized that following co-exposure, germ cells adapt to the lack of Sertoli cell support by reducing the Fas response to normal FasL signals. These findings provide a better understanding and appreciation of the tissue complexity and technical difficulties associated with examining gene expression in the testis.

Suppression of radiation-induced testicular germ cell apoptosis by 2,5-hexanedione pretreatment. I. Histopathological analysis reveals stage dependence of attenuated apoptosis.

In the testis, developing germ cells are dependent on supportive physical and paracrine interactions with Sertoli cells. The intimate nature of this relationship is demonstrated by the fact that a toxic insult compromising the stability of Sertoli cells will have deleterious effects on the associated germ cells. 2,5-Hexanedione (HD) and x-radiation (x-ray) are testicular toxicants, each with a unique cellular target. HD exposure disrupts microtubule function in Sertoli cells, and x-ray exposure causes double-strand breaks in the DNA of germ cells. Despite their differing modes of action, exposure to either toxicant has the similar ultimate effect of increased germ cell apoptosis. In this study, adult male F344 rats were exposed to 1% HD in the drinking water for 18 days with or without coexposure to 2 or 5 Gy x-ray 12 h prior to necropsy. Incidence of retained spermatid heads was increased in the HD and coexposure groups. Germ cell apoptosis was significantly increased in the x-ray and coexposure groups. There was a striking stage-dependent attenuation of apoptosis with coexposure compared with x-ray alone. Detailed histopathological analysis revealed a significant suppression of x-ray-induced germ cell apoptosis by HD pretreatment in stages I-VI of the seminiferous cycle, most noticeably at stages II/III. We hypothesize either that subacute HD pretreatment compromises the ability of the Sertoli cells to eliminate x-ray-damaged germ cells or that germ cells are more resistant to x-ray-induced damage, having adapted to a less supportive environment.

Inguinal hernia containing functioning, rudimentary uterine horn and endometriosis.

BACKGROUND: Inguinal hernia containing uterus and endometriosis is exceedingly rare. Most inguinal endometriosis is located at an extrapelvic site near the round ligament. We report a case of a patient with inguinal hernia containing rudimentary uterine horn and endometriosis. CASE: A young, nulliparous, regularly menstruating woman manifested right inguinal mass and pain in the mass during menstruation. At 20 years old, she underwent a surgical procedure for right inguinal mass. Postoperative pathology findings demonstrated inguinal endometriosis. Based on the findings of magnetic resonance imaging, a history of inguinal endometriosis, and the occurrence of inguinal pain during menstruation, she was diagnosed as having incarcerated inguinal hernia containing anomalous uterus and endometriosis. A functioning, noncommunicating, rudimentary uterine horn and endometriosis were surgically removed from the hernia sac. Laparoscopy demonstrated intraabdominal unicornuate uterus, but no pelvic endometriosis. CONCLUSION: Functioning, incarcerated hernia uterus inguinale may be associated with müllerian abnormality and concomitant occurrence of inguinal endometriosis.

Theoretical investigation of the electronic asymmetry of the special pair cation radical in the photosynthetic type-II reaction center.

The electronic asymmetry of the special pair cation radical in the photosynthetic reaction center was studied, using density functional calculations with a polarizable continuum model and a point charge model as the protein environment. The calculated spin density distribution between the halves of the special pair from Rhodobacter sphaeroides agreed well with the experimental value due to the protein polarity effect. The differences in the specific orientations of the ester carbonyl groups of the phytyl groups, as well as the methyl ester groups, are one of the origins of the electronic asymmetry. The orientations of these groups are specific, as revealed by the 14 recent X-ray structures of a variety of type-II reaction centers, with a few exceptions. In addition to the spin density distributions of the special pair cation radical from other species, the structural features of the protein surrounding these groups of a variety of type-II reaction centers were investigated to elucidate the generality and mechanisms of the specific orientations of the groups by a structural alignment and a multiple sequence alignment. The determining factors of the electronic asymmetry among type-II reaction centers are discussed.

Spermatid head retention as a marker of 2,5-hexanedione-induced testicular toxicity in the rat.

Histopathological analysis is a basic methodology for assessing testicular injury after exposure to candidate therapeutics or toxicants. One possible injury response in rat testis is the failure of step 19 spermatids to spermiate. Such spermatids are transported toward the basement membrane, where they are retained for degradation by Sertoli cells. In control rats, these retained spermatid heads (RSH) were observed at Stages IX-XII. Exposure to the Sertoli cell toxicant, 2,5-hexanedione (HD), for eighteen days at 0.08%-1.0% in drinking water resulted in a dose-dependent increase in the number of RSH at Stages IX-XII (no observed effect level [NOEL], 0.14%). To explore the dynamics of spermatid head retention, rats were treated with 0.33% or 1% HD for various durations and RSH were assessed across all stages. After 0.33% HD exposure for eighteen days, there were more RSH present in Stage IX-XII tubules compared to control. Numbers of RSH dropped back to control levels after four weeks of recovery after the eighteen-day exposure. Exposure of rats to 1% HD for eighteen days resulted in markedly elevated numbers of RSH at Stages IX-II/III. There was no evidence of other histopathological alterations. These data identify RSH as a sensitive histopathological marker of testicular toxicity for subacute HD exposure.

A leiomyomatoid angiomatous neuroendocrine tumor of the myometrium: case study with ultrastructural analysis.

Leiomyomatoid angiomatous neuroendocrine tumor (LANT) is a possible new disease entity that was described as a dimorphic neurosecretory tumor with a leiomyomatous vascular component; it was found in the pituitary. We describe here a second case of LANT in a 45-year-old woman with a myometrial tumor, diagnosed clinically as uterine leiomyoma. She underwent laparoscopic myomectomy. The tumor consisted of hyalinized vasculature, containing factor VIII-positive endothelium and smooth muscle actin-positive vascular smooth muscle cells, and stromal cells, expressing neuroadhesion molecules. Both vascular and stromal components diffusely expressed chromogranin A and, as evidenced by electron microscopy, possessed smooth muscle actin filaments and electron-dense neurosecretory granules, which contained the neurosecretory hormone somatostatin. Although no cytokeratin-positive cells were observed, some tumor cells had positive Grimelius staining for argyrophilic granules. These findings meet the definition of LANT, and the occurrence of our case suggests that LANT is a special type of neuroendocrine neoplasm and is not organ specific.

Drug-metabolizing enzyme and transporter expression in a mouse model of diabetes and obesity.

Obesity and type II diabetes pose a serious human health risk. Obese or diabetic patients usually take prescription drugs that require hepatic and renal metabolism and transport, and these patients sometimes display different pharmacokinetics of these drugs. Therefore, mRNA and protein expression of drug-metabolizing enzymes (DMEs) and transporters was measured in livers and kidneys of adult wild-type and ob/ob mice, which model obesity and diabetes. mRNA expression of numerous DMEs increased by at least 2-fold in livers of male ob/ob mice, including Cyp4a14, Cyp2b10, NAD(P)H:quinone oxidoreductase 1 (Nqo1), and sulfotransferase 2a1/2. In general, expression of uptake transporters was decreased in livers of ob/ob mice, namely organic anion-transporting polypeptides (Oatps) and sodium/taurocholate cotransporting polypeptide (Ntcp). In particular, Oatp1a1 mRNA and protein expression in livers of ob/ob mice was diminished to <5% and <15% of that in wild-types, respectively. Generally, the mRNA and protein expression of efflux transporters multidrug resistance-associated proteins (Mrps) was increased in livers of ob/ob mice, particularly with Mrp4 expression being elevated by at least 6-fold and Mrp2 expression at least 3-fold in livers of ob/ob mice. In kidney, Nqo1, Mrp3, 4, Oatp1a1, and organic anion transporter 2 (Oat2) showed significant alterations with mRNA expression levels in ob/ob mice, being increased for Nqo1 and Mrp4 and decreased for Mrp3, Oatp1a1, and Oat2. In summary, the expression of a number of DMEs and transporters was significantly altered in livers and kidneys of ob/ob mice. Since expression of some DMEs and transporters is regulated similarly between mouse and human, the data from this study suggest that transporter expression in liver and kidney may be changed in patients presenting with obesity and/or type II diabetes.

Zinc finger domain of Snail functions as a nuclear localization signal for importin beta-mediated nuclear import pathway.

Snail, a DNA-binding zinc finger protein, functions as a transcriptional repressor for genes including E-cadherin during development and the acquisition of tumor cell invasiveness. Human Snail is a 264-amino acid nuclear protein with an amino-terminal basic amino acid-rich domain (SNAG domain) and a carboxyl-terminal DNA-binding domain (zinc finger domain). A series of fusion proteins composed of green fluorescent protein (GFP) and portions of the Snail protein were generated, and their subcellular localization was examined. Fusion of the four zinc fingers to GFP led to the targeting of GFP to the nucleus, demonstrating that the zinc finger domain is sufficient for nuclear localization. Using an in vitro transport system, the nuclear import of Snail was reconstituted by importin (karyopherin) beta in the presence of Ran and NTF2. We further demonstrated that Snail binds directly to importin beta in a zinc finger domain-dependent manner. These results indicate that zinc finger domain of Snail functions as a nuclear localization signal and Snail can be transported into the nucleus in an importin beta-mediated manner.

Body fat patterning in polycystic ovary syndrome women as a predictor of the response to clomiphene.

BACKGROUND: To investigate the difference in the response to clomiphene citrate (CC) based on body fat distribution in women with polycystic ovary syndrome (PCOS). METHODS: Ninety anovulatory PCOS women were divided into two subgroups based on treatment response: women who ovulated with CC (CC responders, n = 49) and those who did not ovulate with CC (CC nonresponders, n = 41). Baseline characteristics included age, age at menarche, height, weight and body mass index [BMI; weight/(height)2]. Percentage of body fat, body fat mass and the ratio of trunk fat to leg fat mass amount (trunk-leg fat ratio) were measured by dual-energy X-ray absorptiometry (DEXA). RESULTS: Age, age at menarche and height did not differ between the two groups. However, trunk-leg fat ratio in CC responders (0.9 +/- 0.4) was significantly lower than that in CC nonresponders (1.3 +/- 0.4) (p < 0.001). Percentage of body fat, body fat mass and BMI were also lower in CC responders (p < 0.01). On multiple regression analysis, however, trunk-leg fat ratio proved to be a superior predictor of CC responder to percentage of body fat, BMI or body fat mass (standardized regression coefficient > or = 0.510; t-values > or = 3.432; p < 0.001). CONCLUSIONS: Response to CC in anovulatory PCOS women differs with body fat distribution.

Changes in serum leptin levels during GnRH agonist therapy.

The purpose of the present study was to investigate changes in serum leptin levels during GnRH agonist therapy. Twenty regularly menstruating women with uterine leiomyomas were enrolled. These subjects were given GnRH agonist (leuprorelin acetate, 3.75 mg) monthly for 4 months. Serum leptin and estradiol (E2) levels were measured at the two time points of day 1 or 2 of the menstrual cycle and the end of GnRH agonist therapy. Weight, total body fat mass, percentage of body fat, and total body lean mass were measured by whole body scanning with dual-energy X-ray absorptiometry. The ratio of serum leptin levels to total body fat mass (leptin-fat mass ratio), and the ratio of serum leptin levels to total body lean mass (leptin-lean mass ratio) were calculated. All subjects became amenorrheic after the initial administration of GnRH agonist. Baseline E2 levels were 45.4 +/- 21.0 pg/mL, which significantly decreased after GnRH agonist therapy (13.3 +/- 4.2 pg/mL, p<0.01). Baseline leptin levels were 8.7 +/- 8.1 ng/mL, which did not differ from the values after 4 months of GnRH agonist administration (8.9 +/- 6.8 ng/mL). Total body fat mass significantly increased from 20.0 +/- 10.4 to 21.0 +/- 9.4 kg (p<0.05), while total body lean mass significantly decreased (34.5 +/- 4.2 kg to 33.3 +/- 3.9 kg, p<0.01). However, leptin-fat mass ratio after GnRH agonist therapy did not differ from the baseline values (0.39 +/- 0.16 ng/mL/kg vs 0.38 +/- 0.16 ng/mL/kg). Hypogonadism does not have a major impact on circulating leptin levels.

Changes in anthropometry with testosterone therapy in a female with gender identity disorder.

A 31-year-old regularly menstruating Japanese female was referred to our outpatient clinic by a psychiatrist. She had been diagnosed as having gender identity disorder by detailed counseling and clinical intervention 3 years earlier. After obtaining fully informed written consent, we treated her with 125 mg of testosterone enanthate, intramuscularly, every 2 weeks for 4 months. Serum testosterone levels increased to the normal male value (from 28 to 432 ng/dL). Although menstrual cycle remained regular, her voice became lower after 4 months of therapy. Body weight, body mass index, and lean body mass increased, while body fat mass and percentage of body fat decreased. However, trunk-leg fat ratio did not change during the observation period. During testosterone therapy, a disproportionate increase in lean body mass and decrease in body fat mass are early onset events, while the shift toward upper body fat distribution may be a late onset event along with increase in BMD.

Inverse relationship between the changes in trunk lean and fat mass during gonadotropin-releasing hormone agonist therapy.

OBJECTIVE: The aim of the present study was to investigate the relationship between the changes in lean and fat mass during gonadotropin-releasing hormone agonist (GnRH agonist) therapy. METHODS: Subjects were 24 premenopausal women (mean age, 39.5+/-9.4 years; range, 32-52 years) with uterine leiomyomas. They were given GnRH agonist (leuprorelin acetate, 3.75 mg) monthly for 4 months. Age and height were recorded. Body weight, regional and total body composition, and the ratio of trunk fat mass to leg fat mass (trunk-leg fat ratio) were assessed by whole body scanning with dual-energy X-ray absorptiometry. Changes in these variables were investigated. Relationships between the changes in regional lean and fat mass were investigated using Pearson's correlation test. RESULTS: Trunk fat mass significantly increased from 8616+/-3538 to 9265+/-3526 g (P<0.01) and trunk-leg fat ratio significantly increased (1.02+/-0.39 to 1.07+/-0.39, P<0.05). Trunk lean mass significantly decreased from 18,509+/-2602 to 17,916+/-2402 g (P<0.01). However, body weight, and lean and fat mass component in the extremities did not change. Change in trunk fat mass was inversely correlated with change in trunk lean mass (r=-0.439, P<0.05), but such relationships were not observed in arm and leg regions. CONCLUSION: Inverse relationship between the changes in trunk lean and fat mass is observed during GnRH agonist therapy.