Based on recent studies showing that phospholipase D (PLD)1 is associated with intracellular membranes and promotes membrane budding from the trans-Golgi, we tested its possible role in the membrane trafficking of GLUT4 glucose transporters. Using immunofluorescence confocal microscopy, expressed Myc epitopetagged PLD1 was found to associate with intracellular vesicular structures by a mechanism that requires its N-terminal pleckstrin homology domain. Partial co-localization with expressed GLUT4 fused to green fluorescent protein in both 3T3-L1 adipocytes and Chinese hamster ovary cells was evident. Furthermore, microinjection of purified PLD into cultured adipocytes markedly potentiated the effect of a submaximal concentration of insulin to stimulate GLUT4 translocation to cell surface membranes. Insulin stimulated PLD activity in cells expressing high levels of insulin receptors but no such insulin effect was detected in 3T3-L1 adipocytes. Taken together, these results are consistent with the hypothesis that PLD1 associated with GLUT4-containing membranes acts in a constitutive manner to promote the mechanism of GLUT4 translocation by insulin.Physiological glucose homeostasis in humans is largely dependent on the actions of the hormone insulin, particularly its ability to enhance glucose transport into fat and muscle. Insulin exerts this effect primarily through a process whereby sequestered, intracellular GLUT4 glucose transporter proteins are rapidly redistributed to cell surface membranes where they can catalyze glucose uptake into cells (1-3). In both the basal and insulin-stimulated states, GLUT4 proteins appear to cycle between intracellular membrane and plasma membrane locations, but exocytosis is relatively slow in the absence of insulin (4). The action of insulin to stimulate this process appears dependent on the activity of p85/p110-type phosphatidylinositol (PI) 1 3-kinase because it is blocked by inhibitors of this enzyme (5, 6) or by expression of dominant inhibitory constructs of the p85 regulatory subunit (7). Insulin receptor tyrosine kinase activity activates PI 3-kinase through recruitment of Src homology 2 domains of the p85 subunit to phosphotyrosine sites on insulin receptor substrate proteins (8). The concept that the 3Ј-polyphosphoinositide products of insulin-stimulated PI 3-kinase are key mediators of insulin action on GLUT4 translocation is consistent with extensive data supporting a major role of these lipids in membrane trafficking (9).Much effort has been expended attempting to identify downstream targets of the PI 3-kinase products, phosphatidylinositol 3-phosphate (PtdIns(3)P), PtdIns(3,4)P 2 , and PtdIns(3,4,5)P 3 , that might connect insulin receptor signaling to GLUT4 glucose transporter trafficking (1). Some evidence (10, 11) has implicated Akt/ protein kinase B, which is thought to be activated in the presence of 3Ј-polyphosphoinositide through the actions of PDK1 and another unknown protein kinase (12), in GLUT4 regulation. However, other data do not support this view but i...

Conflict of interest: A patent application (30435.354-US-P2) has been filed by BP, FAP, and AD on a method for generation of therapeutic pericytes from human pluripotent stem cells.

Massive tears of the rotator cuff (RC) are often associated with progressive and irreversible muscle degeneration due to fibrosis, fatty infiltration, and muscle atrophy. RC tears are common in individuals older than 60 years and the repair of these tears is amongst the most prevalent of orthopedic procedures. However, most current models of this injury are established in young animals, which may not accurately recapitulate the clinical condition. In this study, we used a murine model of massive RC tears to evaluate agerelated muscle degeneration following chronic injury. The expression of the fibro-adipogenic genes encoding collagen type III and leptin was higher in aged RC compared with matched injured young tissue at 2 weeks post-injury, and development of fibrosis was accelerated in aged mice within 5 days post-injury. Furthermore, the synthesis of collagens type I and III and fat tissue accumulation were significantly higher in injured RCs of aged mice. Similar frequency of fibro-adipogenic PDGFRβ + PDGFRα + progenitor cells was measured in noninjured RC of aged and young mice, but PDGFRβ + PDGFRα + cells contributed to significantly larger fibrotic lesions in aged RCs within 2 weeks post-injury, implying a more robust fibrotic environment in the aged injured muscle. Altogether, these findings demonstrate agedependent differences in RC response to chronic injury with a more profound fibro-adipogenic change in aged muscles. Clinically, cell therapies for muscular pathologies should not only consider the cell type being transplanted but also the recipient milieu into which these cells are seeded.

Background: Individuals with genetic susceptibility to breast cancer may pursue bilateral prophylactic mastectomy (BPM) and subsequent breast reconstruction. This study aimed to characterize immediate reconstructive trends following BPM. Methods: The ACS-NSQIP database (2010 -2019) was used to examine differences in demographics and operative outcomes based on breast reconstruction technique following BPM and factors predicting reconstruction type. Results: Of 1945 patients (mean age, 43.8 ± 11.3 years), implant-based reconstruction (IBR) was most frequently (71.8%) performed following BPM. Patients who underwent IBR (n = 1396) were younger (42.6 years, P < 0.001), more likely to be White (P < 0.05), and more likely to have a BMI less than 25 (P < 0.001). Patients who underwent autologous reconstruction (AR) (n = 186, 45.8 years) were more likely to be Black or African American and have a BMI of 25–30. Patients who underwent mastectomy only (MO) without immediate reconstruction (n = 363) were older (47.6 years), more likely to be Asian, and more likely to have a BMI greater than 35. The MO cohort had the highest frequency of diabetes or smoking history. AR was associated with longer operations, longer lengths of stay, and increased complications. Increasing age and BMI were predictive of AR or MO compared to IBR. Smoking was predictive of MO. Conclusion: This is the first large-scale study of genetically susceptible patients who underwent BPM demonstrating a significant relationship between patient demographics, operative outcomes, and immediate reconstruction technique. These results provide valuable insight for surgeons and patients during the shared decision-making process.

Small animal models of massive tears of the rotator cuff (RC) were introduced a decade ago and have been extensively used to study the pathophysiology of chronically injured RC. Transection of rodent suprascapular nerve and RC tendon results in progressive muscle atrophy, fibrosis and fat accumulation and affect the infraspinatus and supraspinatus muscles similarly to that seen in the setting