Sex-biased islet β cell dysfunction is caused by the MODY MAFA S64F variant by inducing premature aging and senescence in males

Abstract

<jats:title>Summary</jats:title><jats:p>A heterozygous missense variant in the islet β-cell-enriched MAFA (Ser(S)64Phe(F)) transcription factor was identified in humans who developed either diabetes or insulinomatosis, with males more prone to diabetes. This mutation engenders increased stability to the normally unstable MAFA protein. To obtain insight into how this impacts β cell function, we developed a mouse model expressing S64F MafA and found sex-dependent phenotypes, with heterozygous mutant males displaying impaired glucose tolerance while females were slightly hypoglycemic and had improved blood glucose clearance. Only heterozygous males showed higher MafA protein levels, which preceded the onset of glucose intolerance and sex dependent variations in expression of genes involved in aging, DNA damage, calcium signaling, and senescence. Changes in islet calcium handling and signs of aging and senescence processes were validated in male animals. Together, these results implicate accelerated islet aging and senescence in promoting diabetes in male human S64F MAFA carriers.</jats:p>

Lipid Droplets Protect Human β-Cells From Lipotoxicity-Induced Stress and Cell Identity Changes

“…EndoCβH2 cells were propagated in DMEM (Gibco and Thermo Fisher Scientific, Waltham, MA) in presence of 5.6 mmol/L glucose, 2% BSA (Serologicals Proteins, Kankakee, IL), 100 μU/mL penicillin, 100 μg/mL streptomycin, 50 μmol/L 2-mercaptoethanol, 10 mmol/L nicotinamide, 5 μg/mL transferrin, and 6.7 ng/mL sodium selenite (Sigma-Aldrich, St. Louis, MO) as previously described ( 15 , 16 ). Lentiviral vectors encoding shRNAs that were either scrambled (shScramble) (i.e., Sham) or to PLIN2 (shPLIN2) were constructed by VectorBuilder (Chicago, IL), as was the PLIN2 protein overexpressing cassette, with each containing a puromycin selection marker.…”

“…cDNA libraries were constructed and paired-end sequencing was performed on an Illumina NovaSeq 6000 (150 nucleotide reads). The generated FASTQ files were processed and interpreted with the Genialis visual informatics platform ( https://www.genialis.com ) ( 15 ). Sequence quality checks were performed with use of raw and trimmed reads with FastQC ( https://www.bioinformatics.babraham.ac.uk/projects/fastqc ), and Trimmomatic was used to trim adapters and filter out poor-quality reads.…”

Lipid droplets protect human β cells from lipotoxic-induced stress and cell identity changes

“…EndoCβH2 cells were propagated in Dulbecco’s Modified Eagle Medium (Gibco, Thermo Fisher) in presence of 5.6mM glucose, 2% BSA (Serologicals Proteins Inc., Kankakee, IL), 100 μU/ml penicillin, 100 μg/ml streptomycin, 50 μM 2-mercaptoethanol, 10 mM nicotinamide, 5 μg/ml transferrin and 6.7 ng/ml sodium selenite (Sigma-Aldrich) as described previously (15,16). Lentiviral vectors encoding short hairpin RNAs that were either Scrambled (i.e., Sham) or to PLIN2 (shPLIN2) were constructed by VectorBuilder (Chicago, IL) as was the PLIN2 protein over-expressing cassette, with each containing a puromycin selection marker.…”

“…EndoCβH2 cells were propagated in Dulbecco's Modified Eagle Medium (Gibco, Thermo Fisher) in presence of 5.6mM glucose, 2% BSA (Serologicals Proteins Inc., Kankakee, IL), 100 μU/ml penicillin, 100 μg/ml streptomycin, 50 μM 2-mercaptoethanol, 10 mM nicotinamide, 5 μg/ml transferrin and 6.7 ng/ml sodium selenite (Sigma-Aldrich) as described previously (15,16).…”

“…cDNA libraries were constructed and paired-end sequencing performed on an Illumina NovaSeq6000 (150 nucleotide reads). The generated FASTQ files were processed and interpreted using the Genialis visual informatics platform (https://www.genialis.com) with previously described process (15). Sequence quality checks were determined using raw and trimmed reads with FastQC (http://www.bioinformatics.babraham.ac.uk/projects/fastqc), and Trimmomatic was utilized to trim adapters and filter out poor-quality reads.…”