A reappraisal of ovarian stimulation strategies used in assisted reproductive technology.

Ovarian stimulation is a fundamental step in assisted reproductive technology (ART) with the intention of inducing ovarian follicle development prior to timed intercourse or intra-uterine insemination and facilitating the retrieval of multiple oocytes during a single in vitro fertilization (IVF) cycle. The basis of ovarian stimulation includes the administration of exogenous gonadotropins, with or without pre-treatment with oral hormonal therapy. Gonadotropin-releasing hormone agonist or antagonist is given in addition to the gonadotropins to prevent a premature rise of endogenous luteinizing hormone that would in turn lead to premature ovulation. With the advancement in technology, various stimulation protocols have been devised to cater for different patient needs. However, ovarian hyperstimulation syndrome and its serious complications may occur following ovarian stimulation. It is also evident that suboptimal ovarian stimulation strategies may have a negative impact on oogenesis, embryo quality, endometrial receptivity, and reproductive outcomes over recent years. This review describes the various forms of pre-treatment for ovarian stimulation and stimulation protocols, and aims to provide clinicians with the latest available evidence.

Hyperandrogenism, Elevated 17-Hydroxyprogesterone and Its Urinary Metabolites in a Young Woman with Ovarian Steroid Cell Tumor, Not Otherwise Specified: Case Report and Review of the Literature.

We describe a case of a 24-year-old overweight woman who presented with hirsutism, secondary amenorrhea, clitoromegaly, and symptoms of diabetes mellitus (DM). While a diagnosis of polycystic ovary syndrome (PCOS) with its associated metabolic disturbances was initially considered, serum total testosterone, androstenedione, and 17-hydroxyprogesterone (17-OHP) measured by liquid chromatography tandem mass spectrometry (LC-MS/MS) were significantly increased. As 17-OHP did not increase upon ACTH (Synacthen) stimulation and the urinary steroid profile (USP) was compatible with an ovarian source of 17-OHP excess rather than adrenal, non classical congenital adrenal hyperplasia (NCCAH) was unlikely and an androgen-secreting tumor was suspected. Transabdominal ultrasound revealed the presence of an enlarged right ovary with a polycystic ovary morphology and no discrete mass. Transvaginal ultrasound and [18F]- fluorodeoxyglucose positron emission tomography-computed tomography (FDG PET-CT) enabled the localization of a right ovarian tumor. Laparoscopic right salpingo-oophorectomy was performed and a histological diagnosis of steroid cell tumor, not otherwise specified (SCT-NOS) was made. Hyperandrogenism and menstrual disturbances resolved postoperatively. A literature review revealed that 17-OHP-secreting SCT-NOS may uncommonly show positive responses to ACTH stimulation similar to 21-hydroxylase deficiency. Alternatively, USP might be useful in localizing the source of 17-OHP to the ovaries. Its diagnostic performance should be evaluated in further studies.

Efficacy and outcomes of transobturator tension-free vaginal tape with or without concomitant pelvic floor repair surgery for urinary stress incontinence: five-year follow-up.

OBJECTIVES: To compare the 5-year subjective and objective outcomes of transobturator tension-free vaginal tape alone versus the same procedure with concomitant pelvic floor repair surgery for pelvic organ prolapse in women with urinary stress incontinence. DESIGN: Prospective cohort study. SETTING: Urogynaecology unit at a university hospital in Hong Kong. PATIENTS: Of 218 women, 96 (44%) received transobturator tension-free vaginal tape alone and 122 (56%) received transobturator tension-free vaginal tape with concomitant pelvic floor repair surgery from September 2004 to December 2009. The women were followed up annually for up to 5 years after the operation. MAIN OUTCOME MEASURES: The 5-year subjective and objective cure rates were assessed. Subjective cure was defined as no urine loss during physical activity and objective cure was defined as no urine leakage on coughing during urodynamic study. RESULTS: Overall, 88 women receiving transobturator tension-free vaginal tape alone and 101 women receiving transobturator tension-free vaginal tape with concomitant pelvic floor repair surgery were followed up for 5 years after operation. The subjective and objective cure rates of the two groups were 70.5% versus 94.1% (P<0.01) and 80.3% versus 85.7% (P=0.58), respectively. CONCLUSIONS: Transobturator tension-free vaginal tape is an effective treatment for urinary stress incontinence in women who received it alone or with concomitant pelvic floor repair surgery for pelvic organ prolapse, providing high subjective and objective efficacy for up to 5 years after operation. Transobturator tension-free vaginal tape with concomitant pelvic floor repair surgery achieved similar, if not better, long-term outcome compared with transobturator tension-free vaginal tape alone.

Immunology in the clinic review series: focus on type 1 diabetes and viruses: the role of viruses in type 1 diabetes: a difficult dilemma.

Convincing evidence now indicates that viruses are associated with type 1 diabetes (T1D) development and progression. Human enteroviruses (HEV) have emerged as prime suspects, based on detection frequencies around clinical onset in patients and their ability to rapidly hyperglycaemia trigger in the non-obese diabetic (NOD) mouse. Whether or not HEV can truly cause islet autoimmunity or, rather, act by accelerating ongoing insulitis remains a matter of debate. In view of the disease's globally rising incidence it is hypothesized that improved hygiene standards may reduce the immune system's ability to appropriately respond to viral infections. Arguments in favour of and against viral infections as major aetiological factors in T1D will be discussed in conjunction with potential pathological scenarios. More profound insights into the intricate relationship between viruses and their autoimmunity-prone host may lead ultimately to opportunities for early intervention through immune modulation or vaccination.

The CVB and etiology of type 1 diabetes.

The origins of type 1 diabetes (T1D) are largely unknown. Fewer than 50% of the cases of the disease are attributable to host genetics, indicating that environmental factors are involved in disease development. The most often cited environmental agents implicated as initiators of T1D are the human enteroviruses, in particular the group B coxsackieviruses (CVB). Although the connection between the CVB and T1D has not been firmly established, significant' evidence supports the role of these pathogens in T1D development.

The clinical achievement portfolio: an outcomes-based assessment project in nursing education.

Dynamic healthcare market forces impel educators to search for innovative methods of academic assessment to measure learning outcomes. The clinical achievement portfolio is a creative and systematic tool for documenting continuous improvement of student clinical learning. The authors describe the use of the portfolio as a pilot project aimed at introducing reflective thinking and measuring clinical learning in undergraduate nursing education. Potential benefits of the clinical portfolio and implications for future research are proposed.

Expression of the coxsackievirus and adenovirus receptor in cultured human umbilical vein endothelial cells: regulation in response to cell density.

Primary cultures of human umbilical vein endothelial cells (HUVEC) express the human coxsackievirus and adenovirus receptor (HCAR). Whereas HCAR expression in HeLa cells was constant with respect to cell density, HCAR expression in HUVEC increased with culture confluence. HCAR expression in HUVEC was not quantitatively altered by infection with coxsackievirus B.

Purification of the putative coxsackievirus B receptor from HeLa cells.

We have identified a protein expressed by human and murine cells susceptible to coxsackievirus B3 (CVB3) infection and purified it from HeLa cells. This protein of approximately 45,000 Mr is expressed by HeLa cells and mouse fetal heart fibroblasts (susceptible to infection), and not by C3H murine fibroblasts or the human RD cell line (resistant). The protein was isolated from Triton X-100- deoxycholate lysates of HeLa cells by chromatography on concanavalin A-Sepharose, Affi-gel Blue, Phenyl Sepharose, and PBE94. The CVB3-binding fraction from PBE94 was blotted from SDS-polyacrylamide gel onto PVDF membrane for amino acid sequencing. Approximately 2 pmoles of CVB3-binding protein provided assignments for 26 consecutive residues: LSITTPEEMIEKAKGETAYLPXKFTL. This sequence corresponds neither to decay accelerating factor nor to nucleolin, both of which have previously been identified as CVB3-binding proteins, but does match two entries in GenBank. These data show that we have purified a novel CVB3-binding protein, the characteristics of which suggest the CVB group receptor has been purified. Identification of 26 amino acid residues in the protein and corresponding GenBank enteries will accelerate study of CVB tropism and the diseases caused by these viruses.

Coxsackievirus-induced chronic myocarditis in murine models.

Challenge of several murine strains with two highly myocarditic variants of coxsackievirus B3 (CVB3) induced acute and chronic myocarditis, detectable at 21 and 45 days post-inoculation (p.i.). In-situ hybridization of coronal heart sections showing chronic inflammation with a radiolabelled CVB3 probe detected viral genomic RNA at day 7 p.i. but rarely at 21 or 45 days p.i., suggesting few murine heart cells actively replicate virus during chronic myocardial inflammation. Data will be presented that favour an alternative hypothesis, i.e. autoimmune responses to shared epitopes among CVB3 proteins, cardiac myosin and myocardial cell surface proteins (molecular mimicry) can affect the severity of chronic inflammation. Mice inoculated with human cardiac myosin (HM) prior to a CVB3m challenge develop less myocarditis than mice inoculated with virus only, suggesting that antibodies stimulated by HM bind virus, reduce the virus burden and provide protection. Mice inoculated with HM only develop non-neutralizing antibodies against purified CVB3m particles. Several strains of mice inoculated with specific synthetic peptides of HM produce antibodies against CVB3m and/or develop cardiomyopathy. Thus antigen-challenged mice can produce antibodies which cross-react among CVB3m HM or cardiac cells to protect or exacerbate heart disease.

Tissue factor and the extrinsic pathway of coagulation during infection and vascular inflammation.

Tissue factor is a potent initiator of blood coagulation. In tissue sections, it has been immunologically demonstrated in cells normally not in contact with circulating blood, and elevated activity has been repeatedly demonstrated in peripheral blood monocytes of patients considered to be at risk for thrombosis. Studies with endothelial cells and monocytes in culture have documented the induction of tissue factor synthesis by biochemical mediators of the inflammatory process. Lytic processes, such as those caused by complement activation or viral infections, increase the tissue factor activity several fold over the basal level of the affected cells. Diminished anti-thrombotic properties of endothelium, and induced tissue factor expression in endothelium and monocytes/macrophages, combined with the increased specific procoagulant activity resulting from cell membrane damage, may endow inflammatory foci with dramatically elevated procoagulant activity. Levels of tissue factor activity at which procoagulant mechanisms escape regulation by natural anticoagulant mechanisms and produce thrombosis remain to be determined.

Progress and challenges in the pathological diagnosis of myocarditis.

The pathological diagnosis of myocarditis rests on well-described histopathological criteria. Appreciation for the disease-specific sensitivity of the endomyocardial biopsy, as well as the phenotypical and functional nature of inflammatory infiltrates, will enhance the clinical utility of the biopsy technique. New information regarding the role of enteroviruses in immunological sensitization and the study of enteroviruses in murine models and human patients are fostering new perspectives and routes of investigation of appropriate therapeutic interventions for myocarditis.

Cap-binding complex protein p220 is not cleaved during echovirus 22 replication in HeLa cells.

Previously we demonstrated that echovirus 22 is an atypical enterovirus which does not shut off host cell protein synthesis. We extend these findings by showing that echovirus 22 does not cleave p220, part of the cellular cap-binding complex necessary for cap-dependent translation, suggesting a biology more consistent with cardioviruses than enteroviruses.

Evidence for a group-specific enteroviral antigen(s) recognized by human T cells.

Human peripheral blood mononuclear cells from 15 normal, healthy adult volunteers proliferated in vitro against a panel of enteroviral antigens, including coxsackievirus B3, coxsackievirus B2, coxsackievirus B6, coxsackievirus A16, and poliovirus 1. No proliferation against the cardiovirus encephalomyocarditis virus occurred. Lymphocytes obtained from cord blood drawn from seven neonates were uniformly nonresponsive to enteroviral antigens. Although serum neutralization antibody titers indicated different exposure histories of the volunteers, only one had a titer against coxsackievirus B6, a rare isolate in the United States. The peripheral blood mononuclear cells from each volunteer responded in vitro to each enterovirus tested even though not all individuals had serum neutralizing antibody against each virus. The predominant cell type responding in vitro was the CD4+ T cell. Denaturation of viral antigen by Formalin did not prevent the recognition of the common group antigen by the T cells, indicating that noninfectious virus can also serve as antigen. These data demonstrate that human T cells recognize a common enterovirus group antigen(s).

Echovirus 22 is an atypical enterovirus.

Although echovirus 22 (EV22) is classified as an enterovirus in the family Picornaviridae, it is atypical of the enterovirus paradigm, typified by the polioviruses and the coxsackie B viruses. cDNA reverse transcribed from coxsackievirus B3 (CVB3) RNA does not hybridize to genomic RNA of EV22, and conversely, cDNA made to EV22 does not hybridize to CVB3 genomic RNA or to molecular clones of CVB3 or poliovirus type 1. EV22 cDNA does not hybridize to viral RNA of encephalomyocarditis virus or to a molecular clone of Theiler's murine encephalomyelitis virus, members of the cardiovirus genus. The genomic RNA of EV22 cannot be detected by the polymerase chain reaction using generic enteroviral primers. EV22 does not shut off host cell protein synthesis, and the RNA of EV22 is efficiently translated in vitro in rabbit reticulocyte lysates. Murine enterovirus-immune T cells recognize and proliferate against EV22 as an antigen in vitro, demonstrating that EV22 shares an epitope(s) common to enteroviruses but not found among other picornaviruses.

Murine cell-mediated immune response recognizes an enterovirus group-specific antigen(s).

Splenocytes taken from mice inoculated with coxsackievirus B3 (CVB3) (Nancy) developed an in vitro proliferative response against CVB3 antigen. This response could not be detected earlier than 8 days postinoculation but could be detected up to 28 days after exposure to CB3. CVB3-sensitized splenocytes responded not only to the CVB3 antigen but to other enteroviruses as well. This response was found to be enterovirus specific in that no response was detected to a non-enteroviral picornavirus, encephalomyocarditis virus, or to an unrelated influenza virus. The generation of a splenocyte population capable of responding to an enterovirus group antigen(s) was not limited to inoculation of mice with CVB3, as similar responses were generated when mice were inoculated with CVB2. Cell subset depletions revealed that the major cell type responding to the enterovirus group antigen(s) was the CD4+ T cell. Current evidence suggests that the group antigen(s) resides in the structural proteins of the virus, since spleen cells from mice inoculated with a UV-inactivated, highly purified preparation of CVB3 virions also responded in vitro against enteroviral antigens.

Factors affecting the detection of enteroviruses in cerebrospinal fluid with coxsackievirus B3 and poliovirus 1 cDNA probes.

Enteroviruses are common pathogens of meningitis and encephalitis, and infections are often difficult to distinguish clinically from bacterial and herpetic infections of the central nervous system. An array of enteroviruses added to cerebrospinal fluid in reconstruction experiments were detected by a dot hybridization assay. Optimal handling and processing conditions for infected cerebrospinal fluid were established, and the effect on the hybridization reaction of humoral and cellular components of the inflammatory response was determined. Six hybridization probes, derived from poliovirus 1 and coxsackievirus B3, were then tested, singly and in combinations, to optimize the sensitivity and spectrum of the assay. Implications for enteroviral taxonomy based on these experiments are discussed.

Virus detection by nucleic acid hybridization: examination of normal and ALS tissues for the presence of poliovirus.

A nucleic acid hybridization assay was developed as a sensitive assay for the presence of poliovirus RNA in human tissue. The assay could detect the presence of an average of one poliovirus per 200 cells. A method for determining the extent of degradation of the tissue RNA was developed and used to show that a significant fraction of human central nervous system (CNS) autopsy material contains highly degraded RNA which is unsuitable for hybridization studies. A total of 15 different control and amyotrophic lateral sclerosis tissues were assayed for the presence of poliovirus-like RNA. Virus RNA was detected in one of the control tissues and in none of the ALS tissues.