R.P. Gada

Comparison and validation of point of care lactate meters as a replacement for fetal pH measurement

OBJECTIVE To validate a point of care lactate device to replace fetal pH measurement. STUDY DESIGN AND METHODS Cord blood samples drawn immediately following delivery were tested on the Nova Lactate Plus and ARKRAY Lactate Pro, the Corometrics 220 pH System, and the Vitros chemistry analyzer (used as lactate reference). RESULTS Nova demonstrated a constant positive bias relative to the lactate reference method; while the Lactate Pro correlated well with the reference method up to 6 mmol/L. Receiver operating characteristic (ROC) curve analysis showed optimal sensitivity and specificity for predicting pH<7.20 at lactate values of 6.8 mmol/L for the Nova and 4.8 mmol/L for the Lactate Pro. CONCLUSION Using Lactate Pro the best cut-off for predicting pH< or =7.20 was 4.8 mM; which coincides with current clinical cut-offs. Thus any lactate device that correlates well with the laboratory reference method can be used with a clinical cut-off of 4.8 mmol/L.

A Novel Role of the Sp/KLF Transcription Factor KLF11 in Arresting Progression of Endometriosis

Endometriosis affects approximately 10% of young, reproductive-aged women. Disease associated pelvic pain; infertility and sexual dysfunction have a significant adverse clinical, social and financial impact. As precise disease etiology has remained elusive, current therapeutic strategies are empiric, unfocused and often unsatisfactory. Lack of a suitable genetic model has impaired further translational research in the field. In this study, we evaluated the role of the Sp/KLF transcription factor KLF11/Klf11 in the pathogenesis of endometriosis. KLF11, a human disease-associated gene is etiologically implicated in diabetes, uterine fibroids and cancer. We found that KLF11 expression was diminished in human endometriosis implants and further investigated its pathogenic role in Klf11-/- knockout mice with surgically induced endometriotic lesions. Lesions in Klf11-/- animals were large and associated with prolific fibrotic adhesions resembling advanced human disease in contrast to wildtype controls. To determine phenotype-specificity, endometriosis was also generated in Klf9-/- animals. Unlike in Klf11-/- mice, lesions in Klf9-/- animals were neither large, nor associated with a significant fibrotic response. KLF11 also bound to specific elements located in the promoter regions of key fibrosis-related genes from the Collagen, MMP and TGF-β families in endometrial stromal cells. KLF11 binding resulted in transcriptional repression of these genes. In summary, we identify a novel pathogenic role for KLF11 in preventing de novo disease-associated fibrosis in endometriosis. Our model validates in vivo the phenotypic consequences of dysregulated Klf11 signaling. Additionally, it provides a robust means not only for further detailed mechanistic investigation but also the ability to test any emergent translational ramifications thereof, so as to expand the scope and capability for treatment of endometriosis.

Detailed structural-functional analysis of the Kruppel-like factor 16 (KLF16) transcription factor reveals novel mechanisms for silencing Sp/KLF sites involved in metabolism and endocrinology.

Background: KLF16 is the least characterized family member of recently described metabolic regulators. Results: We extensively characterize mechanisms of DNA binding and chromatin coupling used by KLF16 to regulate metabolic gene expression. Conclusion: KLF16 is a novel regulator of metabolic genes by regulatable coupling to Sin3-histone deacetylase complexes. Significance: This knowledge reveals key mechanisms used by KLF16 as a regulator of metabolic gene expression. Krüppel-like factor (KLF) proteins have elicited significant attention due to their emerging key role in metabolic and endocrine diseases. Here, we extend this knowledge through the biochemical characterization of KLF16, unveiling novel mechanisms regulating expression of genes involved in reproductive endocrinology. We found that KLF16 selectively binds three distinct KLF-binding sites (GC, CA, and BTE boxes). KLF16 also regulated the expression of several genes essential for metabolic and endocrine processes in sex steroid-sensitive uterine cells. Mechanistically, we determined that KLF16 possesses an activation domain that couples to histone acetyltransferase-mediated pathways, as well as a repression domain that interacts with the histone deacetylase chromatin-remodeling system via all three Sin3 isoforms, suggesting a higher level of plasticity in chromatin cofactor selection. Molecular modeling combined with molecular dynamic simulations of the Sin3a-KLF16 complex revealed important insights into how this interaction occurs at an atomic resolution level, predicting that phosphorylation of Tyr-10 may modulate KLF16 function. Phosphorylation of KLF16 was confirmed by in vivo 32P incorporation and controlled by a Y10F site-directed mutant. Inhibition of Src-type tyrosine kinase signaling as well as the nonphosphorylatable Y10F mutation disrupted KLF16-mediated gene silencing, demonstrating that its function is regulatable rather than constitutive. Subcellular localization studies revealed that signal-induced nuclear translocation and euchromatic compartmentalization constitute an additional mechanism for regulating KLF16 function. Thus, this study lends insights on key biochemical mechanisms for regulating KLF sites involved in reproductive biology. These data also contribute to the new functional information that is applicable to understanding KLF16 and other highly related KLF proteins.

Unilateral oophorectomy results in compensatory follicular recruitment in the remaining ovary at time of ovarian stimulation for in vitro fertilization.

OBJECTIVE To assess the effect of unilateral oophorectomy (UO) by assessing ovarian reserve (OVR) and the response to gonadotropin stimulation in women with UO undergoing in vitro fertilization (IVF) compared with the response of the ipsilateral ovary of women without UO. DESIGN Historical cohort study. SETTING Academic fertility clinic. PATIENT(S) Fifty-one women with single ovary compared with a referent group with both ovaries in a 1:2 fashion. INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Day-3 follicle-stimulating hormone (FSH), estradiol, and antral follicle counts as measures of OVR, and IVF outcomes including number of follicles aspirated and oocytes retrieved. RESULT(S) The baseline demographics and serum markers of OVR were not different. Referent women had greater follicular yield and oocyte numbers when compared with women with UO; however, when compared with the ipsilateral ovary of the referents, women with UO had a higher antral follicle count and greater follicle and oocyte numbers. In multivariate analyses, the ovary from women with UO was more likely to yield more than the median number of follicles and oocytes than the ipsilateral ovary in referent women. Live-birth rates in both groups were similar. CONCLUSION(S) Our results suggest that the remaining ovary appears to compensate in follicular yield after UO in women, confirming the animal data. Women with UO can be reassured and appropriately counseled regarding IVF.

Potential of inner cell mass outgrowth and amino acid turnover as markers of quality in the in vitro fertilization laboratory

OBJECTIVE To compare sensitivity of inner cell mass (ICM) outgrowth assay and analysis of culture media amino acid turnover with the sensitivity of the human sperm motility assay (HSMA) and murine embryo assay (MEA) for detection of formaldehyde toxicity. DESIGN Prospective in vitro study. SETTING University hospital-based infertility center. ANIMAL(S) Murine embryos. INTERVENTION(S) The HSMA, MEA, and ICM outgrowth assays were performed with media containing 0-64-μM concentrations of formaldehyde. These assays were compared with dynamics of amino acid turnover in culture media. MAIN OUTCOME MEASURE(S) The lowest concentration of formaldehyde in culture media detected by each quality control assay. RESULT(S) Sperm forward progression, but not motility, detected formaldehyde at a concentration of 32 μM. Sperm motility index identified formaldehyde toxicity at 64 μM, whereas blastocyst rates in the MEA were affected at 32 μM formaldehyde. Evaluation of ICM using outgrowth and grade detected 16 μM formaldehyde. Leucine turnover in culture media detected 64 μM formaldehyde in the amino acid assay. CONCLUSION(S) Inner cell mass outgrowth is a more sensitive bioassay than MEA and HSMA for the detection of formaldehyde in culture media. Amino acid metabolism may also provide a sensitive quality control measure for detection of formaldehyde.

A Hemangioma of the Cervix in Childhood Can Be a Harbinger of Menorrhagia and Infertility as an Adult

Early diagnosis of uterine hemangiomas may direct management decisions improving long-term outcome in children, adolescents and adulthood.