E.F. Wolff

Endometrial stem cell transplantation restores dopamine production in a Parkinson’s disease model

Parkinson’s disease (PD) is a neurodegenerative disorder caused by the loss of dopaminergic neurons. Adult human endometrial derived stem cells (HEDSC), a readily obtainable type of mesenchymal stem‐like cell, were used to generate dopaminergic cells and for transplantation. Cells expressing CD90, platelet derived growth factor (PDGF)‐Rβ and CD146 but not CD45 or CD31 were differentiated in vitro into dopaminergic neurons that exhibited axon projections, pyramidal cell bodies and dendritic projections that recapitulate synapse formation; these cells also expressed the neural marker nestin and tyrosine hydroxylase, the rate‐limiting enzyme in dopamine synthesis. Whole cell patch clamp recording identified G‐protein coupled inwardly rectifying potassium current 2 channels characteristic of central neurons. A 1‐methyl 4‐phenyl 1,2,3,6‐tetrahydro pyridine induced animal model of PD was used to demonstrate the ability of labelled HEDSC to engraft, migrate to the site of lesion, differentiate in vivo and significantly increase striatal dopamine and dopamine metabolite concentrations. HEDSC are a highly inducible source of allogenic stem cells that rescue dopamine concentrations in an immunocompetent PD mouse model.

Demonstration of Multipotent Stem Cells in the Adult Human Endometrium by In Vitro Chondrogenesis

Stem cells are defined by their unique capacity for self-renewal and multilineage differentiation. Stem cells have been obtained from multiple extramedullary tissues. Recently, a population of progenitor cells have been identified in the endometrium. However, multilineage differentiation of endometrial stem cells has not been reported.Endometrial tissue was obtained from reproductive-aged women undergoing surgery for benign disease, from which monolayer endometrial stromal cell (ESC), myometrial, fibroid, fallopian tube, and uterosacral ligament tissue cultures were generated. Once confluent, cells were trypsinized and centrifuged in conical tubes to form a cell pellet. Cell pellets were cultured in a defined chondrogenic media (CM) containing dexamethasone and transforming growth factor (TGF)—β2 or TGF-β3 for 3 to 21 days. Samples were analyzed for markers of human articular cartilage, including sulfated glycosaminoglycans and expression of type II collagen. ESC pellets cultured in CM were found to contain cells that resemble chondrocytes. These cells expressed sulfated glycosaminoglycans and type II collagen typical of human articular cartilage. Myometrial, fibroid, fallopian tube, and uterosacral ligament cells were unable to undergo chondrogenic differentiation using the pellet culture method. Cells derived from the endometrium were able to differentiate into a heterologous cell type: chondrocytes, thus demonstrating the presence of multipotent stem cells. Endometrium is a potential source of multipotent stem cells.

Derivation of Insulin Producing Cells From Human Endometrial Stromal Stem Cells and Use in the Treatment of Murine Diabetes

Pancreatic islet cell transplantation is an effective approach to treat type 1 diabetes, however the shortage of cadaveric donors and limitations due to rejection require alternative solutions. Multipotent cells derived from the uterine endometrium have the ability to differentiate into mesodermal and ectodermal cellular lineages, suggesting the existence of mesenchymal stem cells in this tissue. We differentiated human endometrial stromal stem cells (ESSC) into insulin secreting cells using a simple and nontransfection protocol. An in vitro protocol was developed and evaluated by assessing the expression of pan β-cell markers, followed by confirmation of insulin secretion. PAX4, PDX1, GLUT2, and insulin, were all increased in differentiated cells compared to controls. Differentiated cells secreted insulin in a glucose responsive manner. In a murine model, differentiated cells were injected into the kidney capsules of diabetic mice and human insulin identified in serum. Within 5 weeks blood glucose levels were stabilized in animals transplanted with differentiated cells, however those treated with undifferentiated cells developed progressive hyperglycemia. Mice transplanted with control cells lost weight and developed cataracts while those receiving insulin producing cells did not. Endometrium provides an easily accessible, renewable, and immunologically identical source of stem cells with potential therapeutic applications in diabetes.

Self-reported menopausal symptoms, coronary artery calcification, and carotid intima-media thickness in recently menopausal women screened for the Kronos early estrogen prevention study (KEEPS)

OBJECTIVE To determine whether self-reported menopausal symptoms are associated with measures of subclinical atherosclerosis. DESIGN Cross-sectional analysis. SETTING Multicenter, randomized controlled trial. PATIENT(S) Recently menopausal women (n = 868) screened for the Kronos Early Estrogen Prevention Study (KEEPS). INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Baseline menopausal symptoms (hot flashes, dyspareunia, vaginal dryness, night sweats, palpitations, mood swings, depression, insomnia, irritability), serum E2 levels, and measures of atherosclerosis were assessed. Atherosclerosis was quantified using coronary artery calcium (CAC) Agatston scores (n = 771) and carotid intima-media thickness (CIMT). Logistic regression model of menopausal symptoms and E2 was used to predict CAC. Linear regression model of menopausal symptoms and E2 was used to predict CIMT. Correlation between length of time in menopause with menopausal symptoms, E2, CAC, and CIMT were assessed. RESULT(S) In early menopausal women screened for KEEPS, neither E2 nor climacteric symptoms predicted the extent of subclinical atherosclerosis. Palpitations and depression approached significance as predictors of CAC. Other symptoms of insomnia, irritability, dyspareunia, hot flashes, mood swings, night sweats, and vaginal dryness were not associated with CAC. Women with significantly elevated CAC scores were excluded from further participation in KEEPS; in women meeting inclusion criteria, neither baseline menopausal symptoms nor E2 predicted CIMT. Years since menopause onset correlated with CIMT, dyspareunia, vaginal dryness, and E2. CONCLUSION(S) Self-reported symptoms in recently menopausal women are not strong predictors of subclinical atherosclerosis. Continued follow-up of this population will be performed to determine whether baseline or persistent symptoms in the early menopause are associated with progression of cardiovascular disease. CLINICAL TRIAL REGISTRATION NUMBER NCT00154180.

Endometrial stem cell transplantation in MPTP‐ exposed primates: an alternative cell source for treatment of Parkinson's disease

Parkinsons disease (PD) is a neurodegenerative disease caused by the loss of dopaminergic neurons in the substantia nigra. Cell‐replacement therapies have emerged as a promising strategy to slow down or replace neuronal loss. Compared to other stem cell types, endometrium‐derived stem cells (EDSCs) are an attractive source of stem cells for cellular therapies because of their ease of collection and vast differentiation potential. Here we demonstrate that endometrium‐derived stem cells may be transplanted into an MPTP exposed monkey model of PD. After injection into the striatum, endometrium‐derived stem cells engrafted, exhibited neuron‐like morphology, expressed tyrosine hydroxylase (TH) and increased the numbers of TH positive cells on the transplanted side and dopamine metabolite concentrations in vivo. Our results suggest that endometrium‐derived stem cells may provide a therapeutic benefit in the primate model of PD and may be used in stem cell based therapies.

Ovarian reserve and subsequent assisted reproduction outcomes after methotrexate therapy for ectopic pregnancy or pregnancy of unknown location

OBJECTIVE To assess ovarian reserve after methotrexate treatment for ectopic pregnancy or pregnancy of unknown location after assisted reproductive technology (ART). DESIGN Retrospective cohort study. SETTING Large ART practice. PATIENT(S) Women receiving methotrexate or surgery after ART. INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Follicle-stimulating hormone (FSH), antral follicle count (AFC), and oocyte yield compared between women treated with methotrexate or surgery, with secondary outcomes of clinical pregnancy and live birth. RESULT(S) There were 153 patients in the methotrexate group and 36 patients in the surgery group. Neither group demonstrated differences in ovarian reserve or oocyte yield in a comparison of the before and after treatment values. The change in ovarian reserve and oocyte yield after treatment were similar between the two groups. The number of doses of methotrexate was not correlated with changes in ovarian reserve, indicating no dose-dependent effect. Time between treatment and repeat ART was not correlated with outcomes. Live birth in subsequent cycles was similar in the two groups. CONCLUSION(S) Ovarian reserve and subsequent ART cycle outcomes were reassuring after methotrexate or surgical management of ectopic pregnancy. No adverse impact of methotrexate was detected in this large fertility cohort as has been previously described elsewhere.

Characterization of extracellular DDX4- or Ddx4-positive ovarian cells

To the Editor: A few groups1–5 have now reported that ovarian-derived stem cells (OSCs; also known as oogonial stem cells or oocyte precursor cells) have been isolated from adult mouse5 and rat4 ovaries; these cells are able to undergo meiosis after transplantation back into recipient ovaries and give rise to offspring. These cells have also been isolated from human adult ovaries, where they can give rise to oocytes after xenotransplantation2. The marker used to isolate viable cells with germ cell characteristics, DDX4 in humans or Ddx4 in mice and rats (hereafter referred to collectively as DDX4/Ddx4), is controversial because it was historically considered to be exclusively an intracellular protein distributed in the cytoplasm of germ cells. However, DDX4/Ddx4 was reported to have a C-terminal domain that is expressed extracellularly, whereas the N terminus is expressed intracellularly2,3. DDX4/Ddx4 expression was reported in freshly isolated OSCs and after propagation for 18 months (mouse) and 4 months (human) in defined cultures by immunostaining, reverse-transcription PCR (RT-PCR), and fluorescence-activated cell sorting (FACS)2. Here we further characterize the expression of DDX4/Ddx4 in mouse, rhesus macaque and human ovarian cells using a polyclonal antibody specific to DDX4 (ab13840; Abcam, generously provided by Jonathan Tilly and purchased from Abcam). By immunohistochemical analysis of paraffin-embedded tissue, we observed staining for DDX4/Ddx4 in the expected locations in

Skin wrinkles and rigidity in early postmenopausal women vary by race/ethnicity: baseline characteristics of the skin ancillary study of the KEEPS trial

OBJECTIVE To characterize skin wrinkles and rigidity in recently menopausal women. DESIGN Baseline assessment of participants before randomization to study drug. SETTING Multicenter trial, university medical centers. PATIENT(S) Recently menopausal participants enrolled in the Kronos Early Estrogen Prevention Study (KEEPS). INTERVENTION(S) Skin wrinkles were assessed at 11 locations on the face and neck using the Lemperle wrinkle scale. Skin rigidity was assessed at the forehead and cheek using a durometer. MAIN OUTCOME MEASURE(S) Skin wrinkles and rigidity were compared among race/ethnic groups. Skin wrinkles and rigidity were correlated with age, time since menopause, weight, and body mass index (BMI). RESULT(S) In early menopausal women, wrinkles, but not skin rigidity, vary significantly among races, where black women have the lowest wrinkle scores. In white women, chronological age was significantly correlated with worsening skin wrinkles, but not with rigidity. Skin rigidity correlated with increasing length of time since menopause, however, only in the white subgroup. In the combined study group, increasing weight was associated with less skin wrinkling. CONCLUSION(S) Skin characteristics of recently menopausal women are not well studied. Ethnic differences in skin characteristics are widely accepted, but poorly described. In recently menopausal women not using hormone therapy (HT), significant racial differences in skin wrinkling and rigidity exist. Continued study of the KEEPS population will provide evidence of the effects of HT on the skin aging process in early menopausal women.

Longitudinal changes in menopausal symptoms comparing women randomized to low-dose oral conjugated estrogens or transdermal estradiol plus micronized progesterone versus placebo: the Kronos Early Estrogen Prevention Study.

Objective: The objective of the present study was to compare the efficacy of two forms of menopausal hormone therapy in alleviating vasomotor symptoms, insomnia, and irritability in early postmenopausal women during 4 years. Methods: A total of 727 women, aged 42 to 58, within 3 years of their final menstrual period, were randomized to receive oral conjugated estrogens (o-CEE) 0.45 mg (n = 230) or transdermal estradiol (t-E2) 50 &mgr;g (n = 225; both with micronized progesterone 200 mg for 12 d each mo), or placebos (PBOs; n = 275). Menopausal symptoms were recorded at screening and at 6, 12, 24, 36, and 48 months postrandomization. Differences in proportions of women with symptoms at baseline and at each follow-up time point were compared by treatment arm using exact &khgr;2 tests in an intent-to-treat analysis. Differences in treatment effect by race/ethnicity and body mass index were tested using generalized linear mixed effects modeling. Results: Moderate to severe hot flashes (from 44% at baseline to 28.3% for PBO, 7.4% for t-E2, and 4.2% for o-CEE) and night sweats (from 35% at baseline to 19% for PBO, 5.3% for t-E2, and 4.7% for o-CEE) were reduced significantly by 6 months in women randomized to either active hormone compared with PBO (P < 0.001 for both symptoms), with no significant differences between the active treatment arms. Insomnia and irritability decreased from baseline to 6 months postrandomization in all groups. There was an intermittent reduction in insomnia in both active treatment arms versus PBO, with o-CEE being more effective than PBO at 36 and 48 months (P = 0.002 and 0.05) and t-E2 being more effective than PBO at 48 months (P = 0.004). Neither hormone treatment significantly affected irritability compared with PBO. Symptom relief for active treatment versus PBO was not significantly modified by body mass index or race/ethnicity. Conclusions: Recently postmenopausal women had similar and substantial reductions in hot flashes and night sweats with lower-than-conventional doses of oral or transdermal estrogen. These reductions were sustained during 4 years. Insomnia was intermittently reduced compared with PBO for both hormone regimens.

Are intracytoplasmic sperm injection and high serum estradiol compounding risk factors for adverse obstetric outcomes in assisted reproductive technology

OBJECTIVE To evaluate whether intracytoplasmic sperm injection (ICSI) use and E2 on the final day of assisted reproductive technology (ART) stimulation are associated with adverse obstetric complications related to placentation. DESIGN Retrospective cohort study. SETTING Large private ART practice. PATIENT(S) A total of 383 women who underwent ART resulting in a singleton live birth. INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Adverse placental outcomes composed of placenta accreta, placental abruption, placenta previa, intrauterine growth restriction, preeclampsia, gestational hypertension, and small for gestational age infants. RESULT(S) Patients with adverse placental outcomes had higher peak serum E2 levels and were three times more likely to have used ICSI. Adverse placental outcomes were associated with increasing E2 (odds ratio 1.36, 95% confidence interval 1.13-1.65) and ICSI (odds ratio 3.86, 95% confidence interval 1.61-9.27). Adverse outcomes increased when E2 was >3,000 pg/mL and continued to increase in a linear fashion until E2 was >5,000 pg/mL. The association of ICSI with adverse outcomes was independent of male factor infertility. Interaction testing suggested the adverse effect of E2 was primarily seen in ICSI cycles, but not in conventional IVF cycles. Estradiol >5,000 pg/mL was associated with adverse placental events in 36% of all ART cycles and 52% of ICSI cycles. CONCLUSION(S) ICSI and elevated E2 on the day of hCG trigger were associated with adverse obstetric outcomes related to placentation. The finding of a potential interaction of E2 and ICSI with adverse placental events is novel and warrants further investigation.