Expression of Dystrophin Dp71 Splice Variants Is Temporally Regulated During Rodent Brain Development.

Dystrophin Dp71 is the major product of the Duchenne muscular dystrophy (DMD) gene in the brain, and its loss in DMD patients and mouse models leads to cognitive impairments. Dp71 is expressed as a range of proteins generated by alternative splicing of exons 71 to 74 and 78, classified in the main Dp71d and Dp71f groups that contain specific C-terminal ends. However, it is unknown whether each isoform has a specific role in distinct cell types, brain regions, and/or stages of brain development. In the present study, we characterized the expression of Dp71 isoforms during fetal (E10.5, E15.5) and postnatal (P1, P7, P14, P21 and P60) mouse and rat brain development. We finely quantified the expression of several Dp71 transcripts by RT-PCR and cloning assays in samples from whole-brain and distinct brain structures. The following Dp71 transcripts were detected: Dp71d, Dp71d∆71, Dp71d∆74, Dp71d∆71,74, Dp71d∆71-74, Dp71f, Dp71f∆71, Dp71f∆74, Dp71f∆71,74, and Dp71fΔ71-74. We found that the Dp71f isoform is the main transcript expressed at E10.5 (> 80%), while its expression is then progressively reduced and replaced by the expression of isoforms of the Dp71d group from E15.5 to postnatal and adult ages. This major finding was confirmed by third-generation nanopore sequencing. In addition, we found that the level of expression of specific Dp71 isoforms varies as a function of postnatal stages and brain structure. Our results suggest that Dp71 isoforms have different and complementary roles during embryonic and postnatal brain development, likely taking part in a variety of maturation processes in distinct cell types.

Diabetes Mellitus and Pregnancy: An Insight into the Effects on the Epigenome.

Worldwide, diabetes mellitus represents a growing health problem. If it occurs during pregnancy, it can increase the risk of various abnormalities in early and advanced life stages of exposed individuals due to fetal programming occurring in utero. Studies have determined that maternal conditions interfere with the genotypes and phenotypes of offspring. Researchers are now uncovering the mechanisms by which epigenetic alterations caused by diabetes affect the expression of genes and, therefore, the development of various diseases. Among the numerous possible epigenetic changes in this regard, the most studied to date are DNA methylation and hydroxymethylation, as well as histone acetylation and methylation. This review article addresses critical findings in epigenetic studies involving diabetes mellitus, including variations reported in the expression of specific genes and their transgenerational effects.

Improving Cardiovascular Risk in Postmenopausal Women with an (-)-Epicatechin-Based Nutraceutical: A Randomly Assigned, Double-Blind vs. Placebo, Proof-of-Concept Trial.

BACKGROUND: Age-adjusted rates of cardiovascular disease (CVD) are higher in men than in women. CVD risk-factor outcomes are underrecognized, underestimated, and undertreated in women because the clinical expressions in women differ from those of men. There are no universally accepted recommendations on what to do in women when the values of fasting glucose, blood pressure, and lipids are only slightly altered or at borderline values. We reported the positive effects on CVD risk markers using cacao by-products, showing that alternative approaches can be used to prevent cardiovascular disease in women. The objective was to evaluate the changes in lipoprotein subfractions induced by three months of treatment with an epicatechin-enriched cacao supplement. METHODS: A double-blind, placebo-controlled proof-of-concept study was developed to evaluate the effects of 3 months of treatment with an (-)-epicatechin-enriched cacao supplement on lipoprotein subfractions. RESULTS: The usual screening workshop for postmenopausal women could be insufficient and misleading. Assessing the effect of a (-)-epicatechin-enriched cacao supplement employing a lipoprotein subfractionation profile analysis suggests a decrease in cardiovascular risk. CONCLUSIONS: A simple, low-cost, safe (-)-epicatechin-enriched cacao supplement product can improve the cardiovascular risk in postmenopausal women.

Hyperglycemia affects neuronal differentiation and Nestin, FOXO1, and LMO3 mRNA expression of human Wharton's jelly mesenchymal stem cells of children from diabetic mothers.

Pregestational Diabetes Mellitus (PDM) during pregnancy constitutes an unfavorable embryonic and fetal development environment, with a high incidence of congenital malformations (CM). Neural tube defects are the second most common type of CM in children of diabetic mothers (CDM), who also have an elevated risk of developing neurodevelopmental disorders. The mechanisms that lead to these neuronal disorders in CDM are not yet fully understood. The present study aimed to know the effect of hyperglycemia on proliferation, neuronal differentiation percentage, and expression of neuronal differentiation mRNA markers in human umbilical cord Wharton's jelly mesenchymal stem cells (hUCWJMSC) of children from normoglycemic pregnancies (NGP) and PDM. We isolated and characterized hUCWJMSC by flow cytometry, immunofluorescence, RT-PCR and were induced to differentiate into adipocytes, osteocytes, and neurons. Proliferation assays were performed to determine the doubling time, and Nestin, TUBB3, FOXO1, KCNK2, LMO3, and MAP2 mRNA gene expression was assessed by semiquantitative RT-PCR. Hyperglycemia significantly decreased proliferation and neuronal differentiation percentage in NGP and PDM cells treated with 40 mM d-glucose. Nestin mRNA expression decreased under control glycemic conditions, while FOXO1, KCNK2, LMO3, and MAP2 mRNA expression increased during neuronal differentiation in both NGP and PDM cells. On the other hand, under hyperglycemic conditions, Nestin was significantly decreased in cells from NGP but not in cells from PDM, while mRNA expression of FOXO1 and LMO3 was significantly increased in cells from NGP, but not in cells from PDM. We found evidence that maternal PDM, with hyperglycemia in culture, affects the biological properties of fetal cells. All these results could be part of fetal programming.

Markers of oxidative stress in postmenopausal women with metabolic syndrome.

During the postmenopausal period, there are metabolic alterations that predispose individuals to metabolic syndrome (MS), oxidative stress (OS), and the risk of developing cardiovascular diseases. We aimed to compare the concentrations of OS markers in postmenopausal women with and without MS. Malondialdehyde, carbonyl groups, and total antioxidant capacity (TAC) were quantified. We conducted a cross-sectional study: Group 1 (n = 42) included women without MS, and Group 2 (n = 58) comprised women with MS. Participants' age was similar between groups. Glucose, insulin, the homeostasis model assessment of insulin resistance, triglycerides, uric acid, and body mass index were significantly lower in postmenopausal women without MS. OS markers were significantly lower in Group 1 vs. Group 2: malondialdehyde, 31.32 ± 14.93 vs. 40.27 ± 17.62 pmol MDA/mg dry weight (p = .01); protein carbonylation, 6325 ± 1551 vs. 7163 ± 1029 pmol PC/mg protein (p = .0003); and TAC, 1497 ± 297.3 vs. 1619 ± 278.8 pmol Trolox equivalent/mg protein (p = .041). OS markers were significantly higher in postmenopausal women with MS. Impact statementWhat is already known on this subject? Oxidative stress has been implicated in numerous disease processes; however, information on the relationship between oxidative stress and metabolic syndrome among postmenopausal women remains limited.What do the results of this study add? Our results indicate that in postmenopausal Mexican women, oxidative stress markers were significantly lower in those without metabolic syndrome, whereas total antioxidant capacity was higher in those with metabolic syndrome, which could be explained as an antioxidant defense mechanism capable of neutralising excess oxidative damage markers.What are the implications of these findings for clinical practice and/or further research? This study is of interest to a broad audience because it compares the concentrations of oxidative stress markers in postmenopausal women with and without metabolic syndrome. Our study could support intervention with supplements or foods rich in antioxidants as lifestyle modifications in postmenopausal women with metabolic syndrome.

AZFa, AZFb, AZFc and gr/gr Y-chromosome microdeletions in azoospermic and severe oligozoospermic patients, analyzed from a neural network perspective.

AIM: Analysis of male infertility by molecular methods has increased since recognition of genetic risk factors. The AZFa, AZFb, AZFc, and gr/gr regions on the Y-chromosome can cause male infertility. The aim of this study was to determine the prevalence of Y-chromosome microdeletions in these regions in infertile Mexican patients. MATERIAL AND METHODS: We recruited 57 infertile patients with abnormal sperm count (26 azoospermic and 31 oligozoospermic) and 55 individuals with normal sperm count. Analysis of the regions of interest was performed by PCR. RESULTS: 15.8% of infertile patients presented Y-chromosome microdeletions, whereas no deletions were found in the control group. Deletions were observed in all the analyzed regions except in AZFa. Additionally, the neural network model revealed a mild genotype-phenotype correlation between deletion of the sY1191, sY1291 and sY254 markers with oligozoospermia, azoospermia and cryptozoospermia, respectively. CONCLUSIONS: Our data show that AZFb, AZFc, and gr/gr microdeletions are significantly associated with infertility in Mexican population. In addition, the neural network model revealed a discrete genotype-phenotype correlation between specific deletions and a particular abnormality. Our results reinforce the importance of the analysis of AZF regions as part of the clinical approach of infertile men.

OBJETIVO: La utilización de técnicas moleculares para estudiar la infertilidad masculina se ha incrementado desde el reconocimiento de factores genéticos. Las regiones AZFa, AZFb, AZFc, y gr/gr del cromosoma Y son causa de infertilidad masculina. El objetvo de este estudio fue determinar la prevalencia de microdeleciones en estas regiones en pacientes infértiles Mexicanos. MATERIAL Y MÉTODOS: Reclutamos 57 pacientes infértiles con cuentas espermáticas anormales (26 con azoospermia y 31 con oligozoospermia) y 55 individuos con cuentas espermáticas normales. El análisis de las regiones se realizó mediante PCR. RESULTADOS: 15.8% de los pacientes infértiles presentó microdeleciones, no se encontraron microdeleciones en el grupo control. Las microdeleciones fueron observadas en todas las regiones excepto en AZFa. Adicionalmente, el modelo de red neuronal reveló una leve correlación genotipo-fenotipo entre microdeleciones de los marcadores sY1191, Sy1291 y sY254 con oligozoospermia, azoospermia y criptozoospermia, respectivamente. CONCLUSIONES: Nuestros datos muestran que las microdeleciones en AZFb, AZFc, y gr/gr se asocian significativamente con infertilidad en la población Mexicana. Además, el modelo de red neuronal reveló una discreta correlación genotipo-genotipo entre microdeleciones específicas con una anormalidad en particular. Nuestros resultados refuerzan la importancia del análisis de las regiones AZF en el abordaje de la infertilidad masculina.

Embryonic neural stem/progenitor cells as model to characterize dystrophin and dystrophin-associated proteins expression during neuronal or astrocytic differentiation.

Neural stem/progenitor cells (NSPC) are multipotent cells that renew themselves and could differentiate into neurons and macro glia (astrocytes and oligodendrocytes) of the nervous system during embryonic development. Duchenne muscular dystrophy is a severe type of muscular dystrophy caused by mutations in the dmd gene, and one-third of patients cursed with neuro-cognitive impairments. In this data article, we take advantage of the differentiation capacity of NSPC as a model to increase our knowledge in the neuronal and/or astrocytic differentiation and to evaluate the expression of dystrophins and dystrophin-associated proteins. We showed the characterization of undifferentiated and neuron and/or astrocyte differentiated NSPC. In addition, we evaluated the expression and subcellular localization of dystrophins and ß-dystroglycan in undifferentiated NSPC and differentiated to neurons and astrocytes.•Primary culture of NSPC was characterized by the expression of multipotent markers nestin and Sox2.•Neuronal or astrocytic differentiation of NSPC was performed by basic fibroblast growth factor (FGF2) withdrawal, histamine or ciliary neurotrophic factor (CNTF) treatment, and expression of ßIII-tubulin or glial fibrillary acidic protein (GFAP) as differentiation markers for neurons or astrocytes was evaluated.•This study will contribute to the understanding of dystrophins and dystrophin-associated proteins expression and function during neuronal or astrocytic differentiation of NSPC.

Differential expression of Dp71 and Dp40 isoforms in proliferating and differentiated neural stem cells: Identification of Dp40 splicing variants.

Dp71 and Dp40 are the main products of the DMD gene in the central nervous system, and they are developmentally regulated from the early stages of embryonic development to adulthood. To further study the roles of Dp71 and Dp40 during cell proliferation and neural differentiation, we analyzed Dp71/Dp40 isoform expression at the mRNA level by RT-PCR assays to identify alternative splicing (AS) in the isoforms expressed in rat neural stem/progenitor cells (NSPCs) and in differentiated cells (neurons and glia). We found that proliferating NSPCs expressed Dp71d, Dp71dΔ71, Dp71f, Dp71fΔ71, Dp71dΔ74 and Dp40, as well as two Dp40 isoforms: Dp40Δ63,64 and Dp40Δ64-67. In differentiated cells we also found the expression of Dp71d, Dp71dΔ71, Dp71f, Dp71fΔ71 and Dp40. However, the expression frequencies were different in both stages. In addition, in differentiated cells, we found Dp71fΔ71-74, and interestingly, we did not find the expression of Dp71dΔ74 or the newly identified Dp40 isoforms. In this work we show that NSPC differentiation is accompanied by changes in Dp71/Dp40 isoform expression, suggesting different roles for these isoforms in NSPCs proliferation and neuronal differentiation, and we describe, for the first time, alternative splicing of Dp40.

Effect of the Cut-Off Level for Thyroid-Stimulating Hormone on the Prevalence of Subclinical Hypothyroidism among Infertile Mexican Women.

The primary aim of this study was to compare the prevalence of subclinical hypothyroidism (SCH) using two different cut-off levels for TSH values (≥2.5 mIU/L versus ≥4.1 mIU/L). The secondary objective was to analyze the clinical-biochemical characteristics in women with and without SCH. This was a retrospective cross-sectional study. In total, 1496 Mexican women with infertility were included: Group 1, women with TSH levels ranging between 0.3 and 2.49 mIU/L, n = 886; Group 2, women with TSH between 2.5 and 4.09 mIU/L, n = 390; and Group 3, women with TSH ≥4.1 mIU/L n = 220. SCH prevalence was 40.7% (CI 95%: 38.3-43.3%) with TSH cut-off ≥ 2.5 mIU/L, and 14.7% (CI 95%: 12.7-16.5%) with TSH cut-off ≥ 4.1 mIU/L, (p = 0.0001). The prevalence of overweight was higher in Group 2 than in Groups 1 and 3. Thyroid autoimmunity, obesity and insulin resistance were higher in Group 3 than in Group 1 (p < 0.05). No other differences were observed between groups. Conclusions: The prevalence of SCH in our selected patients increased almost three times using a TSH cut-off ≥ 2.5 mIU/L compared with a TSH cut-off ≥ 4.1 mIU/L. Women with TSH ≥4.1 mIU/L compared with TSH cut-off ≤ 2.5 mIU/L more often presented with obesity, thyroid autoimmunity and insulin resistance.

Prevalence of Thyroid Autoimmunity in Women with Recurrent Pregnancy Loss.

Background and objectives: Thyroid autoimmunity (TAI) has been associated with a significantly increased risk of miscarriage in women with recurrent pregnancy loss (RPL). The aim of this study was to determine the prevalence of TAI in women with RPL and compare the clinical characteristics of positive and negative TAI women. Materials and Methods: This is a retrospective cross-sectional study; 203 women with RPL were included. Thyroid profile, anti-thyroid peroxidase (TPO-Ab), and anti-thyroglobulin (TG-Ab) antibodies were measured in all participants. Clinical characteristics and causes of RPL were compared between positive and negative TAI. Results: Prevalence of TAI was 14.8%; prevalence of positive TPO-Ab and TG-Ab was 12.3% and 4.9%, respectively. Women with TAI had significantly higher concentrations of thyrotropin (TSH) compared to women without TAI (4.8 ± 3.8 versus 3.1 ± 1.1, p = 0.001). There was no significant difference in age, the number of gestations, miscarriages, state of antiphospholipid antibodies (aPL), or causes of RPL between women that were TAI-positive versus TAI-negative. Prevalence of positive TAI by cause of RPL was: endocrine 7/25 (28%), genetic 1/5 (20%), autoimmune 1/5 (20%), anatomic 8/55 (14.5%), and unexplained cause 13/112 (11.6%). Conclusions: The prevalence of TAI in women with RPL is 14.8%. Women with an endocrine cause have the highest prevalence of TAI.

Intensive Medical Nutrition Therapy Alone or with Added Metformin to Prevent Gestational Diabetes Mellitus among High-Risk Mexican Women: A Randomized Clinical Trial.

The aim of this study was to examine the efficacy of intensive medical nutrition therapy (MNT) plus metformin in preventing gestational diabetes mellitus (GDM) among high-risk Mexican women. An open-label randomized clinical trial was conducted. Inclusion criteria were pregnant women with three or more GDM risk factors: Latino ethnic group, maternal age >35 years, body mass index >25 kg/m2, insulin resistance, and a history of previous GDM, prediabetes, a macrosomic neonate, polycystic ovarian syndrome, or a first-degree relative with type 2 diabetes. Women before 15 weeks of gestation were assigned to group 1 (n = 45): intensive MNT-plus metformin (850 mg twice/day) or group 2 (n = 45): intensive MNT without metformin. Intensive MNT included individual dietary counseling, with ≤50% of total energy from high carbohydrates. The primary outcome was the GDM incidence according to the International Association of Diabetes Pregnancy Study Groups criteria. There were no significant differences in baseline characteristics and adverse perinatal outcomes between the groups. The GDM incidence was n = 11 (24.4%) in the MNT plus metformin group versus n = 7 (15.5%) in the MNT without metformin group: p = 0.42 (RR: 1.57 [95% CI: 0.67-3.68]). There is no benefit in adding metformin to intensive MNT to prevent GDM among high-risk Mexican women. Clinical trials registration: NCT01675310.

Sexual dimorphism in insulin resistance in a metabolic syndrome rat model.

OBJECTIVE: We assessed the sex-specific differences in the molecular mechanisms of insulin resistance in muscle and adipose tissue, in a MS rat model induced by a high sucrose diet. METHODS: Male, female, and ovariectomized female Wistar rats were randomly distributed in control and high-sucrose diet (HSD) groups, supplemented for 24 weeks with 20% sucrose in the drinking water. At the end, we assessed parameters related to MS, analyzing the effects of the HSD on critical nodes of the insulin signaling pathway in muscle and adipose tissue. RESULTS: At the end of the treatment, HSD groups of both sexes developed obesity, with a 15, 33 and 23% of body weight gain in male, female, and OVX groups respectively, compared with controls; mainly related to hypertrophy of peripancreatic and gonadal adipose tissue. They also developed hypertriglyceridemia, and liver steatosis, with the last being worse in the HSD females. Compared to the control groups, HSD rats had higher IL1B and TNFA levels and insulin resistance. HSD females were more intolerant to glucose than HSD males. Our observations suggest that insulin resistance mechanisms include an increase in phosphorylated AKT(S473) form in HSD male and female groups and a decrease in phosphorylated P70S6K1(T389) in the HSD male groups from peripancreatic adipose tissue. While in gonadal adipose tissue the phosphorylated form of AKT decreased in HSD females, but not in HSD males. Finally, HSD groups showed a reduction in p-AKT levels in gastrocnemius muscle. CONCLUSION: A high-sucrose diet induces MS and insulin resistance with sex-associated differences and in a tissue-specific manner.

Characterization of the expression of dystrophins and dystrophin-associated proteins during embryonic neural stem/progenitor cell differentiation.

Duchenne muscular dystrophy (DMD) is a genetic disease caused by mutations in the dystrophin gene. Dystrophin is required for the organization of a complex consisting of dystroglycans, sarcoglycans, dystrobrevins and syntrophins, known as the dystrophin-associated proteins complex (DAPC). In addition to muscle degeneration, cognitive impairment has been reported in DMD patients. To characterize a suitable model for studying the embryonic cerebral functions of dystrophin, we analyzed the expression patterns of dystrophins/DAPC in undifferentiated and differentiated embryonic neural stem/progenitor cells (NSPC). We found that NSPC express mRNAs for dystrophins Dp427, Dp140, Dp71 and Dp40; ß-dystroglycan; α- and ß-dystrobrevin; α1-, ß1-, ß2- and γ2-syntrophin; and ß-, γ-, δ- and ε-sarcoglycan. Some of these were differentially regulated during neuronal or astrocytic differentiation. Interestingly, the protein expression levels of Dp140, ß-dystroglycan and α2-dystrobrevin were also differentially regulated. Additionally, we found that proliferating NSPC and differentiated neurons and astrocytes show immuno-positive staining for dystrophins and ß-dystroglycan. Our results show that dystrophins and DAPC components are expressed and regulated during the neuronal or astrocytic differentiation of NSPC, suggesting that these proteins may have different roles in the brain development.

Assessment of lamivudine, zidovudine, lopinavir, and ritonavir plasma levels in HIV-positive pregnant women: Drug monitoring application to improve patient safety.

Simultaneous therapeutic drug monitoring (TDM) of combination antiretroviral therapy (cART) is critical during pregnancy in order to improve clinical follow-up, monitor viral load, and patient adherence to treatment.A modified simple and fast ultra-high performance liquid chromatography coupled with tandem mass spectrometry and electrospray ionization (UPLC-ESI-MS/MS) method was developed and validated according to national and international guidelines for the simultaneous determination of lamivudine (LMV), zidovudine (ZDV), lopinavir (LPV), and ritonavir (RTV) concentrations in 100-µL plasma sample of Human Immunodeficiency Virus (HIV)-positive pregnant women. Protein precipitation using 0.1% formic acid in cold acetonitrile was used for sample preparation. The chromatographic separation was achieved with a run-time of 3.0 minutes and 3-µL injection on an ethylene bridged hybrid C18 column (2.1 µm × 50 mm, 1.7 µm), under gradient conditions using acetonitrile and formic acid (0.1%).The chromatographic method was used to analyze 10 plasma samples from 8 HIV pregnant women as a clinical patient routinely follow-up by applying TDM criteria.The protonated precursor/product ion transitions for LMV (230.18/112.08), ZDV (268.22/127.10), LPV (629.55/447.35), and RTV (721.50/296.20) were recorded in multiple-reaction-monitoring (MRM) mode. The calibration curve was linear in the range of 50-3,000, 75-4,500, 250-15,000, and 25-1,500-ng/mL for LMV, ZDV, LPV, and RTV, respectively. The range of accuracy was 97.2% to 100.1% and precision 3.4% to 12.7%. The method showed specificity and matrix effect values of < 15%. Minimum absolute recovery percentages (%CV) were 90.5 (5.4), 90.8 (5.0), 95.4 (3.5), and 93.7 (6.9), for LMV, ZDV, LPV, and RTV, respectively. Drug concentrations in patient samples had high inter-individual variability with %CV of 91.98%, 77.54%, 53.80%, and 92.16% for ZDV, LMV, LPV, and RTV, respectively. Two of the 8 patients showed no adherence due to the absence of Protease Inhibitors (PIs) levels in plasma.This technique demonstrated to be effective in therapeutic drug monitoring and is intended to be used in population pharmacokinetics specifically for HIV-positive pregnant women.

Parameters of Oxidative Stress in Reproductive and Postmenopausal Mexican Women.

In the reproductive phase, women experience cyclic changes in the ovaries and uterus, and hormones regulate these changes. Menopause is the permanent loss of menstruation after 12 months of amenorrhea. Menopause is also linked to a decrease in estrogen production, causing an imbalance in oxidative stress. We aimed to compare the three stages of lipid peroxidation, protein oxidative damage, and total antioxidant capacity (TAC) between reproductive-aged women (RAW) and postmenopausal women (PMW) in Mexico. We carried out a cross-sectional study with 84 women from Mexico City, including 40 RAW and 44 PMW. To determine the oxidative stress of the participants, several markers of lipid damage were measured: dienes conjugates (DC), lipohydroperoxides (LHP), and malondialdehyde (MDA); exposure to protein carbonyl is indicative of oxidative modified proteins, and TAC is indicative of the antioxidant defense system. Biomarkers of oxidative stress were significantly lower in RAW vs. PMW. DC were 1.31 ± 0.65 vs. 1.7 ± 0.51 pmol DC/mg dry weight (p = 0.0032); LHP were 4.95 ± 2.20 vs. 11.30 ± 4.24 pmol LHP/mg dry weight (p < 0.0001); malondialdehyde was 20.37 ± 8.20 vs. 26.10 ± 8.71 pmol MDA/mg dry weight (p = 0.0030); exposure of protein carbonyl was 3954 ± 884 vs. 4552 ± 1445 pmol PC/mg protein (p = 0.042); and TAC was 7244 ± 1512 vs. 8099 ± 1931 pmol Trolox equivalent/mg protein (p = 0.027). PMW display significantly higher oxidative stress markers compared to RAW; likewise, PMW show a higher TAC.

Müllerian Anomalies Prevalence Diagnosed by Hysteroscopy and Laparoscopy in Mexican Infertile Women: Results from a Cohort Study.

BACKGROUND: To evaluate the prevalence of Müllerian anomalies (MAs) in a cohort of infertile Mexican women candidates for infertility treatments (intrauterine insemination or IVF (In vitro fertilization) cycles). METHODS: We performed a retrospective observational study on a cohort of consecutive women, who underwent hysteroscopy and laparoscopy as part of the basic infertility workup from 2002 to 2014, at our center. Our aim was to calculate the prevalence of MAs and each subtype. RESULTS: A total of 4005 women were included in the study. The MA prevalence was 4.4% (95% CI; 3.8-5.1; n = 177). Among women with MAs, the prevalence of different MA types was: septate uterus 54.2% (n = 96), arcuate uterus 15.8% (n = 28), bicornuate uterus 10.7% (n = 19), unicornuate uterus 8.5% (n = 15), didelphys uterus 6.2% (n = 11) and hypoplasia/agenesis 3.4% (n = 6), unclassifiable 1.1% (n = 2). Women with MAs who achieved pregnancy were: 33.3% (n = 59). The MA associated with the highest pregnancy rate was septate uterus after hysteroscopic correction, at 38.5% (37/96). CONCLUSIONS: The prevalence of MAs among infertile Mexican women can be considered as low, but not negligible. The septate uterus is the most common MA in women with infertility.

Perinatal Outcomes in Mexican Women with Untreated Mild Gestational Diabetes Mellitus Diagnosed by the International Association of Diabetes and Pregnancy Study Groups Criteria.

PURPOSE: To compare the risk of adverse perinatal outcomes (APO) between pregnant women with mild gestational diabetes mellitus (GDM) diagnosed by the International Association of Diabetes and Pregnancy Study Groups (IADPSG) criteria, on no specific treatment, versus pregnant women without GDM. PATIENTS AND METHODS: A retrospective cohort study of pregnant women referred to the Instituto Nacional de Perinatología, in Mexico City, for prenatal care and delivery. Eligibility criteria were singleton pregnancy, age >18 years, gestational age 20-28 weeks, and no history of pre-gestational diabetes. The study population was divided into two groups: Group 1, comprising women with mild GDM defined by one abnormal glucose value at the oral glucose tolerance test (OGTT) according to IADPSG criteria [fasting: 5.1-5.2 mmol/L (92-94 mg/dL) or 2h 8.5-8.56 mmol/L (153-154 mg/dL)], who did not receive specific treatment for GDM, and Group 2, comprising women without GDM, matched for maternal age and pre-gestational body mass index (BMI). Women with two or more abnormal OGTT values, pre-gestational diabetes, any chronic disease, or multiple pregnancies were excluded. RESULTS: As many as 282 women were included in each group. There were no significant differences in basal characteristics between groups. APO analysis showed that newborn weight was significantly higher in Group 1 (3042.4±499g) vs Group 2 (2910±565g) p=0.003; conversely, the incidence of large for gestational age (LGA) and macrosomic neonates was similar in both groups (6 vs 5.7% and 2.1 vs 2.2%, respectively). There were no differences in rates of preeclampsia and gestational hypertension, cesarean and preterm delivery, or premature rupture of membranes. A sub-analysis by maternal pre-gestational BMI showed that LGA incidence was significantly higher among babies born to women with pre-gestational BMI ≥30 kg/m2 in both groups. CONCLUSION: The risk of APO was similar among Mexican women with mild untreated GDM diagnosed by IADPSG criteria, compared to pregnant women without GDM. Pre-gestational BMI was an independent risk factor for LGA.

Hyperglycemia differentially affects proliferation, apoptosis, and BNIP3 and p53 mRNA expression of human umbilical cord Wharton's jelly cells from non-diabetic and diabetic pregnancies.

Diabetes in pregnancy constitutes an unfavorable environment for embryonic and fetal development, where the child has a higher risk of perinatal morbidity and mortality, with high incidence of congenital malformations and predisposition to long-term metabolic diseases that increase with a hypercaloric diet. To analyze whether hyperglycemia differentially affects proliferation, apoptosis, and mRNA expression in cells from children of normoglycemic pregnancies (NGPs) and diabetes mellitus pregnancies (DMPs), we used umbilical cord Wharton jelly cells as a research model. Proliferation assays were performed to analyze growth and determine the doubling time, and the rate of apoptosis was determined by flow cytometry-annexin-V assays. AMPK, BNIP3, HIF1α, and p53 mRNA gene expression was assessed by semi-quantitative RT-PCR. We found that hyperglycemia decreased proliferation in a statistically significant manner in NGP cells treated with 40 mM D-glucose and in DMP cells treated with 30 and 40 mM D-glucose. Apoptosis increased in hyperglycemic conditions in NGP and DMP cells. mRNA expression of BNIP3 and p53 was significantly increased in cells from DMPs but not in cells from NGPs. We found evidence that maternal irregular metabolic conditions, like diabetes with hyperglycemia in culture, affect biological properties of fetal cells. These observations could be a constituent of fetal programming.

Dystrophin Dp71 Isoforms Are Differentially Expressed in the Mouse Brain and Retina: Report of New Alternative Splicing and a Novel Nomenclature for Dp71 Isoforms.

Multiple dystrophin Dp71 isoforms have been identified in rats, mice, and humans and in several cell line models. These Dp71 isoforms are produced by the alternative splicing of exons 71 to 74 and 78 and intron 77. Three main groups of Dp71 proteins are defined based on their C-terminal specificities: Dp71d, Dp71f, and Dp71e. Dp71 is highly expressed in the brain and retina; however, the specific isoforms present in these tissues have not been determined to date. In this work, we explored the expression of Dp71 isoforms in the mouse brain and retina using RT-PCR assays followed by the cloning of PCR products into the pGEM-T Easy vector, which was used to transform DH5α cells. Dp71-positive colonies were later analyzed by PCR multiplex and DNA sequencing to determine the alternative splicing. We thus demonstrated the expression of Dp71 transcripts corresponding to Dp71, Dp71a, Dp71c, Dp71b, Dp71ab, Dp71 Δ110, and novel Dp71 isoforms spliced in exon 74; 71 and 74; 71, 73 and 74; and 74 and 78, which we named Dp71d Δ74 , Dp71d Δ71,74 , Dp71d Δ71,73-74 , and Dp71f Δ74 , respectively. Additionally, we demonstrated that the Dp71d group of isoforms is highly expressed in the brain, while the Dp71f group predominates in the retina, at both the cDNA and protein levels. These findings suggest that distinct Dp71 isoforms may play different roles in the brain and retina.

Identification of Dp71 Isoforms Expressed in PC12 Cells: Subcellular Localization and Colocalization with ß-Dystroglycan and α1-Syntrophin.

Several dystrophin Dp71 messenger RNA (mRNA) alternative splice variants have been described. According to the splicing of exon 78 or intron 77, Dp71 proteins are grouped as Dp71d, Dp71f, and Dp71e, and each group has a specific C-terminal end. In this study, we explored the expression of Dp71 isoforms at the complementary DNA (cDNA) level and the subcellular localization of recombinant Myc-Dp71 proteins in PC12 cells. We determined that PC12 cells express Dp71a, Dp71c, Dp71ab, Dp71e, and Dp71ec mRNA splice variants. In undifferentiated and nerve growth factor-differentiated PC12 Tet-ON cells, Dp71a, Dp71ab, and Dp71e were found to localize and colocalize with ß-dystroglycan and α1-syntrophin in the periphery/cytoplasm, while Dp71c and Dp71ec were mainly localized in the cell periphery and showed less colocalization with ß-dystroglycan and α1-syntrophin. The levels of Dp71a, Dp71e, and Dp71ec were increased in the nucleus of differentiated PC12 Tet-ON cells compared to undifferentiated cells. Dp71 isoforms were also localized in neurite extensions and growth cones.