MOESM2 of Scutellarin regulates microglia-mediated TNC1 astrocytic reaction and astrogliosis in cerebral ischemia in the adult rats

Additional file 2: Scutellarin enhanced IL-1β (A1, A2, A3) and iNOS (B1, B2, B3) expression in TNC1 via BV-2-conditioned medium. In TNC1 astrocytes treated with CM, moderate expression of IL-1β and iNOS was detected (A1, B1). The expression was noticeably increased in CM + L (A2, B2) and further enhanced upon incubation with CM + SL for 24 h (A3, B3) with long cytoplasmic processes with expansions projected by TNC1 astrocytes (A3, B3). Scale bars: 20 μm

MOESM5 of Scutellarin regulates microglia-mediated TNC1 astrocytic reaction and astrogliosis in cerebral ischemia in the adult rats

Additional file 5: Showing Notch-1, NICD, HES-1, Nestin, TNF-α, IL-1β and iNOS expression (red) in GFAP (green) reactive astrocytes at 21 d after MCAO (M) and after scutellarin treatment (M + S). Note that the expression is diminished in MCAO but remained more intense with scutellarin treatment. Scale bars: 50 µm. DAPI-blue

Muscle biopsy specimens stained with hematoxylin and eosin.

<p>(A) Normal control (×200). (B) Specimen from patient 11 exhibiting inflammatory cellular infiltration with necrotic and regenerative fibers (×200). (C) Specimen from patient 15 exhibiting mild nuclear transfer and regenerative fibers (×400).</p

Muscle cell nuclear morphology.

<p>A, C, D: ×12000; B, E, F, G, H: ×25000. (A, B) Normal control. (C, D, G) Abnormal nuclear morphology. (C, D, F) Heterochromatin condensation (arrows). (E) Focal loss of nuclear membrane (arrows). (G) Nucleolar hole (arrows). (H) Accumulation of mitochondria around nucleus (arrows).</p

Clinical and neuroradiological findings of the patients.

<p>CK = creatine kinase; ECG = electrocardiogram; EMG = electromyogram; MRI = magnetic resonance imaging; UL = upper limb; LL = lower limb; ND = not done.</p><p>Clinical and neuroradiological findings of the patients.</p

Schematic of the <i>LMNA</i> gene and lamin A protein indicating mutations.

<p>(A) Schematic of the <i>LMNA</i> gene and lamin A protein. Lamin A is encoded by exons 1–12, whereas lamin C terminates at exon 10 with six unique amino acids at its C-terminal. The alternative splice site for lamin A is indicated. Sequence variations affecting the splice donor or acceptor sites that lead to disease are shown for IVS-8. Missense mutations and deletions are indicated on the lamin A protein, where the head, central rod, and tail domain (incorporating the nuclear localization signal [NLS], and Ig-like fold) are indicated. Novel sequence variants are shown above the gene/protein. (B) Illustration of the evolutionary conservation of residues associated with novel missense mutations located in the coding region of <i>LMNA</i>. Sequences were obtained from the online database, Swiss-Prot (<a href="http://expasy.org/sprot/" target="_blank">http://expasy.org/sprot/</a>) and aligned using the Align online tool (<a href="http://www.uniprot.org/align/" target="_blank">http://www.uniprot.org/align/</a>).</p

Phenotype and genotype of patients.

<p>p.r.: Previously reported.</p><p>Phenotype and genotype of patients.</p

Mutational spectrum of Chinese LGMD patients by targeted next-generation sequencing

<div><p>This study aimed to study the diagnostic value of targeted next-generation sequencing (NGS) in limb-girdle muscular dystrophies (LGMDs), and investigate the mutational spectrum of Chinese LGMD patients. We performed targeted NGS covering 420 genes in 180 patients who were consecutively suspected of LGMDs and underwent muscle biopsies from January 2013 to May 2015. The association between genotype and myopathological profiles was analyzed in the genetically confirmed LGMD patients. With targeted NGS, one or more rare variants were detected in 138 patients, of whom 113 had causative mutations, 10 sporadic patients had one pathogenic heterozygous mutation related to a recessive pattern of LGMDs, and 15 had variants of uncertain significance. No disease-causing mutation was found in the remaining 42 patients. Combined with the myopathological findings, we achieved a positive genetic diagnostic rate as 68.3% (123/180). Totally 105 patients were diagnosed as LGMDs with genetic basis. Among these 105 patients, the most common subtypes were LGMD2B in 52 (49.5%), LGMD2A in 26 (24.8%) and LGMD 2D in eight (7.6%), followed by LGMD1B in seven (6.7%), LGMD1E in four (3.8%), LGMD2I in three (2.9%), and LGMD2E, 2F, 2H, 2K, 2L in one patient (1.0%), respectively. Although some characteristic pathological changes may suggest certain LGMD subtypes, both heterogeneous findings in a certain subtype and overlapping presentations among different subtypes were not uncommon. The application of NGS, together with thorough clinical and myopathological evaluation, can substantially improve the molecular diagnostic rate in LGMDs. Confirming the genetic diagnosis in LGMD patients can help improve our understanding of their myopathological changes.</p></div

Lamin A/C mislocation in mutant-transfected HEK 293 cells.

<p>Transfection was performed using (A) GFP, (B) pEGFP-N1-LMNA, (C) pEGFP-N1-LMNA-R48P, (D) pEGFP-N1-LMNA-R249W, (E) pEGFP-N1-LMNA-I373V, or (F) pEGFP-N1-LMNA-I497_E536del. Lamin A/C of the mutants (C–F) is distributed in clusters and is mislocated compared with that of the wild type (B).</p

Histopathological abnormalities in different LGMD subtypes^a.

<p>Histopathological abnormalities in different LGMD subtypes<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0175343#t001fn001" target="_blank">^a</a>.</p