Fig 2: The pedigrees and sequencing data of the two families carrying BSCL2 mutations.(A) The pedigree and the electropherograms of the individuals of the family carrying the BSCL2 p.S90L (c.269C>T) mutation. The proband (III-1) is denoted by an arrow. Filled symbols represent affected members with neuropathy, open symbols indicate unaffected individuals, circles stand for female, and squares stand for male. (B) The pedigree and the electropherogram of the patient carrying the BSCL2 p.R96H (c.287G>A) mutation. (C) The BSCL2 p.S90L and p.R96H mutations reside in an evolutionarily conserved region, as shown by aligning the amino acid sequences of seipin protein orthologs from various species.

Fig 3: In vitro expression of BSCL2 in HEK293 cells.(A) Representative western blot analysis of seipin in the HEK293 cells transfected with plasmids expressing wild-type (WT), S90L, or R96H seipin, or empty vector (vector control). Actin was used as a loading control. Densitometric quantification is shown below. The error bars indicate standard error of the mean (SEM) from 3 independent experiments. IB, immunoblotting. (B) HEK293 cells were transfected with expression plasmids for the WT, S90L, or R96H seipin for 48 hours and subsequently subjected to cycloheximide-chase assays. Representative western blots is shown above. All values are shown as means ± SEM (n = 4). (C) The messenger RNA (mRNA) levels of BSCL2 in the HEK293 cells transfected with WT, S90L, or R96H seipin expression plasmids for 24 hours. The expression levels of BSCL2 were normalized to those of GAPDH and expressed as a fraction of the WT samples, which was set as 100%. The error bars indicate SEM (n = 3). The asterisk indicates statistically significant difference (**, p < 0.01).

Fig 4: Metabolic and mechanical adipose tissue in Ad-B2(-/-)mice. (A) Whole mouse tibiae stained with osmium tetroxide and (B) quantification of cMAT, rMAT, and total MAT by µCT in 16-week-old Ad-B2(+/-) and Ad-B2(-/-) mice (n = 4–5). Representative H&E staining of MAT from caudal vertebrae from the tail (C) and quantification of adipocyte area and frequency of caudal vertebrae MAT (D) of mice described in A. (E) Representative H&E staining images of IFP sections in 12-week-old Ad-B2(+/-) and Ad-B2(-/-) mice. Scale bar represents 100 µm. Quantification of IFP size (F) and adipocytes per section (G) of mice described in E (n = 7). (H) Bscl2 mRNA levels of 12-week-old Ad-B2(+/-) and Ad-B2(-/-) mice in IFP (n = 7). (I) Representative H&E staining images of IFP sections in WT and SKO mice. Scale bar represents 100 µm. All data are presented as the mean ± SEM, *p < 0.05.