Immunofluorescence and Molecular analysis of <i>Calcium ion channel subunits</i> in wBM-hMSC.

<p>(A) Positive control normal Human Skeletal Muscle Myoblast (HSMM). Scale bar = 10 µm (alpha SMA) and 25 µm (SA, Myosin, Myogenin and Desmin). (B) Undifferentiated wBM hMSC analyzed at 28 days. Scale bar = 10 µm (a-SMA), 25 µm (SA, Myosin, and Desmin), 50 (Myogenin). (C) Original gels demonstrating amplification of calcium ion channel subunit transcripts in wBM hMSCs: -RT: control of reverse transcription without RT enzyme; C−: negative control (water); C+: positive control (HSMM); line 1: wBM hMSCs from first passage; line 2: wBM hMSCs from second passage; line 3: wBM hMSCs from fourth passage; β-actin, housekeeping gene.</p

Myogenic differentiation on laminin matrix and <i>L-type Calcium ion channel subunits</i> analysis.

<p>(A) Phase contrast images of laminin cells: presence of some binucleated cells. Scale bar = 25 µm. (B) Immunofluorescence analysis confirmed the presence of a few binucleated structures positive for desmin, SA, myogenin in BM-hMSC cultured on laminin cells. Scale bar = 25 µm. (C) Original gels demonstrating amplification of <i>calcium ion channel subunit</i> transcripts in laminin cells: –RT: control of reverse transcription without RT enzyme; C−: negative control, water; C+: positive control, HSMM; line 1: control wBM-hMSCs; line 2: laminin cells cultured in DMEM-F12 supplemented with 15% FBS; line 3: laminin cells induced to myogenic differentiation with EGF for 7 days.</p

Mouse BM-mMSCs.

<p>(A) Desmin-positive multinucleated myotubes (nuclei labelled in blue with bisbenzimide) derived from the fusion of C2C12 myoblasts. (B) EGFP-MSCs, plated alone, display a fibroblast-like shape. (C–F) In co-culture with C2C12 cells (desmin-positive, labelled in red), MSCs (green) show long cytoplasmatic processes (arrow). (G–J) EGFP-MSCs adhere to desmin-positive myotubes (arrowhead). Scale bar = 50 µm.</p

Integration of BM-hMSCs in the striate muscle.

<p>(A) Bisbenzimide-labelled BM-hMSCs (in blue) appear integrated into desmin-positive striated muscle fibers (in green). (B) At 4 months, several BM-hMSCs are located in close vicinity (arrowheads) to acetylcholine receptors (α-BTX staining, in red) (C). Proliferative profile of transplanted BM-hMSCs at one and (D) four months after transplantation, as revealed by Ki67 immunohistochemistry (in purple). Scale bar = 50 µm.</p

Bisbenzimide-stained BM-hMSCs (in blue) transplanted into rat bulbocavernosus muscle.

<p>(A) 24 hours after transplantation BM-hMSCs appear undifferentiated with a typical round shape (inset in A, scale bar = 500 µm); (B) one month after engraftment many BM-hMSCs with elongated shape are recognizable among muscular fibers (inset in B, scale bar = 100 µm). (C) At 4 months, migration toward muscle fibers is confirmed by elongated appearance of cells occupying peripheral position (inset b), whereas undifferentiated cells are observed in the core of graft (inset a). Scale bar = 500 µm.</p

Cytofluorimetric and morfological analysis of a representative wBM-hMSCs.

<p>(A) Immunophenotypic analysis of wBM hMSCs showing the negativity of haematopoietic markers CD45, CD14, CD34, and the positivity of CD90, CD29, CD73, CD105, CD44. (B) Analyses of growth rate of wBM hMSCs in terms of cumulative PD. The graphic refers to the median cumulative values (1^st passage: median 2.3 - range 1.4–2.6; 2^nd: median 4.5 - range 2.9–5.9; 3^rd: median 6.4 - range 4.9–8.5).</p

Rilpivirine in HIV-1-positive women initiating pregnancy: to switch or not to switch?

International audienceBackgroundSafety data about rilpivirine use during pregnancy remain scarce, and rilpivirine plasma concentrations are reduced during second/third trimesters, with a potential risk of viral breakthroughs. Thus, French guidelines recommend switching to rilpivirine-free combinations (RFCs) during pregnancy.ObjectivesTo describe the characteristics of women initiating pregnancy while on rilpivirine and to compare the outcomes for virologically suppressed subjects continuing rilpivirine until delivery versus switching to an RFC.MethodsIn the ANRS-EPF French Perinatal cohort, we included women on rilpivirine at conception in 2010–18. Pregnancy outcomes were compared between patients continuing versus interrupting rilpivirine. In women with documented viral suppression (<50 copies/mL) before 14 weeks of gestation (WG) while on rilpivirine, we compared the probability of viral rebound (≥50 copies/mL) during pregnancy between subjects continuing rilpivirine versus those switching to RFC.ResultsAmong 247 women included, 88.7% had viral suppression at the beginning of pregnancy. Overall, 184 women (74.5%) switched to an RFC (mostly PI/ritonavir-based regimens) at a median gestational age of 8.0 WG. Plasma HIV-1 RNA nearest delivery was <50 copies/mL in 95.6% of women. Among 69 women with documented viral suppression before 14 WG, the risk of viral rebound was higher when switching to RFCs than when continuing rilpivirine (20.0% versus 0.0%, P = 0.046). Delivery outcomes were similar between groups (overall birth defects, 3.8/100 live births; pregnancy losses, 2.0%; preterm deliveries, 10.6%). No HIV transmission occurred.ConclusionsIn virologically suppressed women initiating pregnancy, continuing rilpivirine was associated with better virological outcome than changing regimen. We did not observe a higher risk of adverse pregnancy outcomes