Enrichment for murine keratinocyte stem cells based on cell surface phenotype

The identification and physical isolation of epithelial stem cells is critical to our understanding of their growth regulation during homeostasis, wound healing, and carcinogenesis. These stem cells remain poorly characterized because of the absence of specific molecular markers that permit us to distinguish them from their progeny, the transit amplifying (TA) cells, which have a more restricted proliferative potential. Cell kinetic analyses have permitted the identification of murine keratinocyte stem cells (KSCs) as slowly cycling cells that retain [(3)H]thymidine ([(3)H]Tdr) label, termed label-retaining cells (LRCs), whereas TA cells are visualized as rapidly cycling cells after a single pulse of [(3)H]Tdr, termed pulse-labeled cells (PLCs). Here, we report on the successful separation of KSCs from TA cells through the combined use of in vivo cell kinetic analysis and fluorescence-activated cell sorting. Specifically, we demonstrate that murine dorsal keratinocytes characterized by their high levels of alpha(6) integrin and low to undetectable expression of the transferrin receptor (CD71) termed alpha(6)(bri)CD71(dim) cells, are enriched for epithelial stem cells because they represent a minor ( approximately 8%) and quiescent subpopulation of small blast-like cells, with a high nuclear:cytoplasmic ratio, containing approximately 70% of label-retaining cells, the latter being a well documented characteristic of stem cells. Conversely, TA cells could be enriched in a phenotypically distinct subpopulation termed alpha(6)(bri)CD71(bri), representing the majority ( approximately 60%) of basal keratinocytes that are actively cycling, and importantly contain approximately 70% of [(3)H]Tdr pulse-labeled cells. Importantly, immunostaining of dorsal skin revealed the presence of CD71(dim) cells in the hair follicle bulge region, a well documented location for KSCs.     

Effect of pirenzepine on aminopyrine uptake by isolated guinea pig parietal cells

The inhibitory action of pirenzepine on acid secretion of isolated guinea pig parietal cells was investigated by the aminopyrine method. Pirenzepine markedly inhibited acid secretion of isolated parietal cells induced by carbachol in a dose-dependent manner but showed no inhibition on acid secretion stimulated by histamine. These results may suggest a direct action of pirenzepine on muscarinic receptors in parietal cells.     

A Damp-and-Push Technique for the Copolymer (Onyx) Embolization of Dural Arteriovenous Fistula

BackgroundCopolymer (Onyx) embolization is an effective treatment for dural arteriovenous fistula (dAVF), however, some dAVFs have multiple, high-flow feeding vessels, resulting in insufficient embolization. For the treatment of such patients, we have developed a novel flow-control technique, the ‘damp-and-push technique’. The purpose of this study was to evaluate the technical efficiency and safety of this technique.MethodsSeven patients who had been diagnosed with intracranial dAVF were treated by transarterial Onyx embolization using the damp-and-push technique between 2016 and 2019. This technique was designed to reduce blood flow to the shunt site using a balloon catheter in the major feeding vessel other than the one injected with Onyx, leading to better Onyx penetration and enabling more controlled embolization of complex dAVFs. Retrospectively collected data were reviewed to assess the occlusion rates and clinical outcomes.ResultsThe dAVF was at a transverse sinus-sigmoid sinus junction in four patients, in the superior sagittal sinus in two, and in the tentorium in one. Five cases were Cognard type Ⅱb and two cases were Cognard type Ⅳ. All the patients were treated by transarterial Onyx injection via the main feeding vessel, combined with flow reduction in the other main feeding vessel using a balloon catheter. Complete occlusion was achieved in six patients and elimination of cerebral venous reflux was achieved in all the patients. There were no immediate or delayed post-interventional complications.ConclusionsTransarterial Onyx embolization of dAVF using the damp-and-push technique is safe and yields a high complete occlusion rate.     

Fine and gross motor skills predict later psychosocial maladaptation and academic achievement

BackgroundDifficulties in fine and gross motor skills are often overlooked as developmental problems, although approximately 6–13% of all school-age children have poor motor coordination. Understanding motor coordination is important from the perspective of school adaptation. This longitudinal cohort study aimed to determine whether fine and gross motor skills in preschool children predict later academic achievement and psychosocial maladaptation.Methods & ProceduresParticipants were 2,501 children from nursery and elementary schools (5–13 years old). The motor skills of preschool children were assessed by their nursery teacher immediately before entering elementary school. The Strengths and Difficulties Questionnaire and a standardized Japanese test were administered annually throughout elementary school.ResultsEarly motor difficulties in preschool children had significant effects on their academic achievement and psychosocial maladaptation up until the sixth grade. Gross motor difficulties in preschool were associated with the later peer problems and worsened emotional symptoms.ConclusionsMotor skills in preschool children are useful as a predictor of later psychosocial maladaptation and academic achievement.     

Dab1‐mediated colocalization of multi‐adaptor protein CIN85 with Reelin receptors,ApoER2 and VLDLR,in neurons

Reelin‐Dab1 signaling is indispensable for proper positioning of neurons in mammalian brain. Reelin is a glycoprotein secreted from Cajal‐Reztuis cells in marginal zone of cerebral cortex, and its receptors are Apolipoprotein E receptor 2 (ApoER2) or very low density lipoprotein receptor (VLDLR) expressed on migrating neurons. When Reelin binds to ApoER2 or VLDLR, an adaptor protein Dab1 bound to the receptors undergoes Tyr phosphorylation that is essential for Reelin signaling. We reported previously that Cdk5‐p35 phosphorylates Dab1 at Ser400 and Ser491 and the phosphorylation regulates its binding to CIN85, which is an SH3‐containing multiadaptor protein involved in endocytic downregulation of receptor‐tyrosine kinases. However, the interaction of CIN85 with Dab1 has not been addressed in neurons. We examined here a possibility that CIN85 has a role in Reelin signaling. We found nonpho‐sphorylated Dab1‐mediated colocalization of CIN85 with ApoER2. The colocalization of CIN85 with ApoER2 was increased in neurons stimulated with Reelin repeats 3‐6, an active Reelin fragment. The stimulation recruited CIN85 to domains in plasma membrane where it colocalized with ApoER2 and Dab1 and then to EEA1‐labeled early endosomes in the cytoplasm. In addition, Tyr phosphorylation of Dab1 strengthened the binding to CIN85. These results suggest that CIN85 participates in Reelin signaling through the binding to Dab1.     

Prediction of motor outcomes and activities of daily living function using diffusion tensor tractography in acute hemiparetic stroke patients

[Purpose] The efficacy of diffusion tensor imaging in the prediction of motor outcomes and activities of daily living function remains unclear. We evaluated the most appropriate diffusion tensor parameters and methodology to determine whether the region of interest- or tractography-based method was more useful for predicting motor outcomes and activities of daily living function in stroke patients. [Subjects and Methods] Diffusion tensor imaging data within 10 days after stroke onset were collected and analyzed for 25 patients. The corticospinal tract was analyzed. Fractional anisotropy, number of fibers, and apparent diffusion coefficient were used as diffusion tensor parameters. Motor outcomes and activities of daily living function were evaluated on the same day as diffusion tensor imaging and at 1 month post-onset. [Results] The fractional anisotropy value of the affected corticospinal tract significantly correlated with the motor outcome and activities of daily living function within 10 days post-onset and at 1 month post-onset. Tthere were no significant correlations between other diffusion tensor parameters and motor outcomes or activities of daily living function. [Conclusion] The fractional anisotropy value of the affected corticospinal tract obtained using the tractography-based method was useful for predicting motor outcomes and activities of daily living function in stroke patients.Key words: Stroke, Diffusion tensor tractography, Activities of daily living function