Regional differences in expression of specific markers for human embryonic stem cells

Characterization of human embryonic stem cell (hESC) lines derived from the inner cell masses of blastocysts generally includes expression analysis of markers such as OCT4, NANOG, SSEA3, SSEA4, TRA-1-60 and TRA-1-81. Expression is usually detected by immunocytochemical staining of entire colonies of hESC, using one colony for each individual marker. Four newly established hESC lines showed the expected expression pattern and were capable of differentiating into the three germ layers in vitro. Neighbouring sections of entire colonies grown for 4, 11, 21 and 28 days respectively were stained with different markers to study the regional distribution and cellular co-expression. TRA-1-60 staining defined the hESC territory at all time points analysed. This territory comprised a characteristic OCT4 and NANOG staining often in overlapping subregions. Staining intensity of nuclei varied from strong OCT4 staining to weak or absent NANOG staining, and vice versa. SSEA4 staining was only observed in small clusters or single cells and not confined to the TRA territory. Co-expression of all markers was only detected in small areas. SSEA1 expression was found exclusively outside the TRA territory. In conclusion, pronounced regional differences in the expression of markers considered specific for undifferentiated hESC may suggest the existence of different cell populations.     

Medical abortion in the first trimester: the use of serum hCG and endometrial thickness as markers of completeness.

OBJECTIVE: The combination of mifepristone and misoprostol is an established method for induction of early first trimester abortion, but there is no consensus about the best evaluation of treatment outcome. We assessed endometrial thickness, determined by ultrasound and serum-human chorionic gonadotropin (s-hCG) as markers of successful management. METHODS: Prospective trial involving 255 women, with a gestation of 62 days or less, who were to undergo medical abortion. In addition to our established routines of performing clinical and ultrasound examinations, we also determined the s-hCG level prior to treatment and at follow-up. RESULTS: Of the 255 subjects treated during the study, 20 (7.8%) were lost to follow-up. The overall complete abortion rate was 94.0%. Fourteen subjects required vacuum aspiration, nine of them prior to the scheduled follow-up and five thereafter. None of the pregnancies were ongoing. A decrease of 99% in s-hCG levels was noted in 99% of the women, when levels determined prior to mifepristone intake and those measured 15-71 days post-abortion were compared. CONCLUSION: This study confirms that s-hCG levels drop sharply after medical abortion. To assess the completeness of medical abortion, we recommend that clinical examination to be combined with determination of s-hCG. Ultrasonography should be carried out only when indicated.     

NCAM in developing mouse gonads and ducts

Summary The expression of theNeuralCell AdhesionMolecule, NCAM, in mouse gonads and ducts was studied from fetal life to maturity. The methods used were immunocytochemical staining and Western blotting. The immunocytochemical studies showed that the only structures that remain NCAM-positive throughout life were the mesonephric-derived rete ovarii and rete testis. Also in the fetal gonads some somatic cell lining the groups of differentiating germ cells were stained. In the immature as well as in the mature ovary the granulosa cells and oocytes of growing and large follicles — but not of small follicles — were stained. A particularly strong staining of the cytoplasm of the oocyte, healthy as well as atretic, was seen. All cells of the testis remained negative except for weakly stained residual bodies and late spermatids. At all ages the male ducts showed only weak staining, whereas in the female Müllerian duct the epithelium became strongly positive at puberty. The stroma of the Müllerian duct was positive during a transitory period around day 16 of fetal life in both sexes. One-dimensional gel immunoblotting of total protein from gonads, rete and ducts from immature and mature mice showed that only the two largest isoforms of NCAM (NCAM-A and NCAM-B) were present. The gonads and the rete of both sexes and the adult uterus expressed only NCAM-B, whereas NCAM-A was also detected in the adult epididymis. The present findings suggest that NCAM may be involved in the normal development and formation of both the gonads and ducts. In particular, NCAM may play a part in sustaining the integrity of the rete testis, thus ensuring the pathway for spermatozoa from the testis to the epididymis. Furthermore this cell adhesion molecule may also be important for follicular growth and differentiation.     

Symptoms as the main problem in primary care: A cross-sectional study of frequency and characteristics

Objective. The aim was to study symptoms managed as the main problem by the general practitioner (GP) and to describe the frequencies and characteristics of presented symptoms when no specific diagnosis could be made. Design. Cross- sectional study. Setting. General practices in the Central Denmark Region. Subjects. In total, 397 GPs included patients with face-to-face contacts during one randomly assigned day in 2008–2009; 7008 patients were included and 5232 presented with a health problem. Main outcome measures. GPs answered a questionnaire after each patient contact. Symptoms and specific diagnoses were subsequently classified using the International Classification of Primary Care (ICPC). Symptom frequency, comorbidity, consultation length, and GP-assessed final outcome and burden of consultations were analysed. Results. The GPs could not establish a specific diagnosis in 36% of patients with health problems. GPs expected that presented symptoms would not result in a future specific diagnosis for half of these patients. Musculoskeletal (lower limb and back) and respiratory (cough) symptoms were most frequent. More GPs had demanding consultations when no specific diagnosis could be made. Higher burden was associated with age, comorbidity, and GP expectancy of persistent symptoms when no diagnosis could be made. Conclusion. Interpretation and management of symptoms is a key task in primary care. As symptoms are highly frequent in general practice, symptoms without a specific diagnosis constitute a challenge to GPs. Nevertheless, symptoms have been given little priority in research. More attention should be directed to evidence-based management of symptoms as a generic phenomenon to ensure improved outcomes in the future.     

Muscle phenotype in patients with myotonic dystrophy type 1

Introduction: The pathogenesis of muscle involvement in patients with myotonic dystrophy type 1 (DM1) is not well understood. In this study, we characterized the muscle phenotype in patients with confirmed DM1. Methods: In 38 patients, muscle strength was tested by hand‐held dynamometry. Myotonia was evaluated by a handgrip test and by analyzing the decrement of the compound muscle action potential. Muscle biopsies were assessed for morphological changes and Na⁺‐K⁺ pump content. Results: Muscle strength correlated with a decline in Na⁺‐K⁺ pump content (r = 0.60, P < 0.001) and with CTG expansion. CTG expansion did not correlate with severity of myotonia, proximal histopathological changes, or Na⁺‐K⁺ pump content. Histopathologically, we found few centrally placed nuclei (range 0.2–6.9%). Conclusions: The main findings of this study are that muscle weakness correlated inversely with CTG expansion and that central nuclei are not a prominent feature of proximal muscles in DM1. Muscle Nerve 47:409‐415, 2013     

Hippocampal involvement in retrieval of odor vs. object memories

The aim of this study was to investigate the role of the human hippocampus in episodic retrieval of odors, in comparison with episodic retrieval of visual objects. Subjects encoded a set of unique odors and objects, and retrieval was tested the next day during functional magnetic resonance imaging (fMRI). Subjects were shown the names of old (studied) and new (unstudied) odors and objects, and asked to indicate which of these stimuli had been presented the previous day. The results showed that brain activation was weaker and more restricted during retrieval of odors than during retrieval of objects, which possibly reflects a general visual dominance effect. Yet, retrieval of odors and objects yielded overlapping clusters of activation the bilateral hippocampi, and the left‐sided activation was specifically increased during successful retrieval (hits > correct rejections) in both modalities. Moreover, retrieval of odors uniquely activated olfactory cortical regions, likely to reflect cortical reinstatement of sensory details. Our fMRI study is the first to make a direct comparison between olfactory and visual episodic memory, and the results provide clear evidence for modality‐independent functions of the hippocampus. © 2012 Wiley Periodicals, Inc.     

Morphometric characteristics of the primordial to primary follicle transition in the human ovary in relation to age

BACKGROUND: The growth pattern of the smallest ovarian follicles in humans is still incompletely documented. Using follicle hemispheres in thick histological sections, morphometric changes of primordial to primary follicles and possible age-related effects were evaluated. METHODS: In ovarian sections from 25 females aged 4-39 years a total of 1122 primordial, transitory or primary follicles were assessed for the diameters of follicles, oocytes and oocyte nuclei and for number of granulosa cells (GCs). RESULTS: The number of GCs in primordial, transitory and primary follicles were approximately 30, 50 and 100, respectively. The diameters of primordial follicles and oocytes increased with the woman's age until the mid-30's, after which time they decreased in size. The number of GCs in primordial follicles showed a moderate increase with age, whereas the number of GCs in transitory follicles showed a clear increase with age . The oocyte nucleus diameter (14-23 microm) showed significant linear correlations with the oocyte and follicular diameters and number of GCs in the follicle, while the number of GCs in the whole follicle and in the largest cross-section were closely related to the oocyte diameter. CONCLUSIONS: The number of GCs in small follicles is accurately estimated and shows an increase with age, indicating that the starting point of follicular development alters with female age. The age-related changes observed may be linked to the poorer reproductive performance of older women.