Differences in blastomere totipotency in 2-cell mouse embryos are a maternal trait mediated by asymmetric mRNA distribution.

It is widely held that the first two blastomeres of mammalian embryos are equally totipotent and that this totipotency belongs to the group of regulative properties. However, this interpretation neglects an important aspect: evidence only came from successful monozygotic twins which can speak only for those pairs of half-embryos that are able to regulate in the first place. Are the frequently occurring incomplete pairs simply an artefact, or do they represent a real difference, be it in the imperfect blastomere's ability to regulate growth or in the distribution of any compound X that constrains regulation? Using the model system of mouse embryos bisected at the 2-cell stage after fertilization, we present evidence that the interblastomere differences evade regulation by external factors and are already latent in oocytes. Specifically, an interblastomere imbalance of epiblast production persists under the most diverse culture conditions and applies to the same extent in parthenogenetic counterparts. As a result, cases in which twin blastocysts continued to develop in only one member account for 65 and 57% of zygotic and parthenogenetic pairs, respectively. The interblastomere imbalance is related to the subcellular distribution of gene products, as documented for the epiblast-related gene Cops3, using mRNA FISH in super-resolution mode confocal microscopy. Blastomere patterns of Cops3 mRNA distribution are α-amanitin-resistant. Thus, the imbalance originates not from de novo transcription, but from influences which are effective before fertilisation. These data expose previously unrecognized limits of regulative capacities of 2-cell stage blastomeres and point to aspects of cytoplasmic organization of the mouse oocyte that segregate unequally to blastomeres during cleavage.

Totipotency segregates between the sister blastomeres of two-cell stage mouse embryos.

Following fertilization in mammals, it is generally accepted that totipotent cells are exclusive to the zygote and to each of the two blastomeres originating from the first mitotic division. This model of totipotency was inferred from a minority of cases in which blastomeres produced monozygotic twins in mice. Was this due to experimental limitation or biological constraint? Here we removed experimental obstacles and achieved reliable quantification of the prevalence of dual totipotency among mouse two-cell stage blastomeres. We separated the blastomeres of 1,252 two-cell embryos, preserving 1,210 of the pairs. Two classes of monozygotic twins became apparent at the blastocyst stage: 27% formed a functional epiblast in both members (concordant), and 73% did so in only one member of the pair (discordant) - a partition that proved insensitive to oocyte quality, sperm-entry point, culture environment and pattern of cleavage. In intact two-cell embryos, the ability of sister blastomeres to generate epiblast was also skewed. Class discovery clustering of the individual blastomeres' and blastocysts' transcriptomes points to an innate origin of concordance and discordance rather than developmental acquisition. Our data place constraints on the commonly accepted idea that totipotency is allocated equally between the two-cell stage blastomeres in mice.

Glenohumeral and scapulothoracic strength impairments exists in patients with subacromial impingement, but these are not reflected in the shoulder pain and disability index.

BACKGROUND: Pain and loss of function are cardinal symptoms associated with Subacromial impingement syndrome (SIS), while the presence and magnitude of deficits in strength and range of motion (ROM) are largely undescribed in non-athletic patients with SIS. Moreover, the relevance of impairments in strength and ROM to patient-reported shoulder function is not well described, even though testing of strength is recommended in clinical guidelines. The purpose of this study was, first, to investigate impairments in glenohumeral and scapulothoracic strength and in abduction and internal rotation ROM in patients with SIS. Secondly, to investigate the influence of these impairments on patient-reported shoulder function. METHODS: Cross-sectional study based on a consecutive cohort of 157 patients referred to specialist examination and diagnosed with shoulder impingement (SIS) using predefined validated diagnostic criteria. Prior to specialist examination, questionnaires regarding shoulder function (Shoulder Pain And Disability Index, SPADI) demographics and kinesiophobia (TSK-11) were collected, and shoulder strength and ROM was measured by trained testers, with the patient reporting pain levels during testing and for the last week. Impairments in strength (abduction, external-rotation, (protraction and horizontal-extension) and ROM (abduction and internal rotation) were investigated in patients with unilateral shoulder pain, using one-sample t-tests. SPADI total score (SPADI) and SPADI function score (SPADI-F), were chosen as dependent variables in multiple regressions to investigate the influence of impairments on patient-reported shoulder function. Independent variables of interest were; strength in abduction and external rotation, abduction ROM, pain-during-tests, pain-last-week and kinesiophobia. RESULTS: Significant impairments were found for all impairment tests, but most pronounced for glenohumeral strength and abduction ROM (29-33% deficits), and less for scapulothoracic strength and internal rotation ROM (8-18% deficits). Pain variables influenced SPADI and SPADI-F score to a high degree (R2 = 23.4-31.6%, p < 0.001), while strength and ROM did not. CONCLUSION: Substantial strength and ROM impairments were found in patients with SIS. Only pain significantly influenced patient-reported function, while impairments did not. As SPADI score does not reflect the substantial strength and ROM impairments in external rotation and abduction observed in patients with SIS, supplemental assessment of these impairments seems important.

Transcription factor cAMP response element modulator (Crem) restrains Pdgf-dependent proliferation of vascular smooth muscle cells in mice.

Transcription factors of the cAMP response element-binding protein (Creb)/cAMP response element modulator (Crem) family were linked to the switch from a contractile to a proliferating phenotype in vascular smooth muscle cells (VSMCs). Here, we analyzed the vascular function of Crem in mice with a global inactivation of Crem (Crem(-/-)). CRE-mediated transcriptional activity was enhanced in primary Crem(-/-) VSMCs under nonstimulated conditions and under stimulation with Forskolin and platelet-derived growth factor (Pdgf) whereas stimulation with nitric oxide or cGMP showed no effect. This elevated CRE-mediated transcriptional activity as a result of Crem inactivation did not alter aortic contractility or fractions of proliferating or apoptotic aortic VSMCs in situ, and no impact of Crem inactivation on the development of atherosclerotic plaques was observed. Crem(-/-) mice exhibited an increased neointima formation after carotid ligation associated with an increased proliferation of VSMCs in the carotid media. Pdgf-stimulated proliferation of primary aortic Crem(-/-) VSMCs was increased along with an upregulation of messenger RNA (mRNA) levels of Pdgf receptor, alpha polypeptide (Pdgfra), cyclophilin A (Ppia), the regulator of G-protein signaling 5 (Rgs5), and Rho GTPase-activating protein 12 (Arhgap12). Taken together, our data reveal the inhibition of Pdgf-stimulated proliferation of VSMCs by repressing the Pdgf-stimulated CRE-mediated transcriptional activation as the predominant function of Crem in mouse vasculature suggesting an important role of Crem in vasculoproliferative diseases.

A practical approach to ultrasonic imaging using diffraction tomography.

A technique for ultrasonic imaging based on the theory of diffraction tomography is presented. The method utilizes a fixed, circular configuration of transmitters and detectors. This configuration was selected because it avoids many practical limitations associated with the design of a medical imaging device. Practical considerations also motivated the inclusion of effects associated with the transmitter beam pattern rather than pursuing the more conventional approach in which plane-wave illumination is required. In addition, the problem of separately imaging both density and compressibility variations is considered.