Draft:Ernesto Bernal-Mizrachi

One Sentence Summary:

Ernesto Bernal-Mizrachi is a physician-scientist at the University of Miami Miller School of Medicine, whose primary research focus is in the factors that promote pancreatic β cell proliferation.

Early Life and Education:

Bernal-Mizrachi is originally from Colombia, and completed his doctorate in medicine (MD) from the University of Valle in the city of Cali, Valle del Cuenca, Colombia in 1989. He continued his medical education with a three-year residency in internal medicine in Miami at the Leonard M. Miller School of Medicine from 1993-1996, and a subsequent three-year fellowship in endocrinology and metabolism at the Washington University School of Medicine in St. Louis from 1996-1999. During this time, Bernal-Mizrachi was mentored by one of the leading figures in the genetics behind diabetes, M. Alan Permutt, who became a professor at the Washington University School of Medicine in St. Louis in 1985 and directed the Diabetes Research and Training Center during Bernal-Mizrachi's fellowship years. That focus on diabetes stemmed from Bernal-Mizrachi's passion to further elucidate the molecular biology behind the chronic ailment and assist in treatment and identification efforts for those in less-developed countries.

Research and Career:

Breakthrough Linkage between Genetics and Diabetes Pathogenesis

One of Bernal-Mizrachi's landmark successes came at the beginnings of his research career. In 1998, Bernal-Mizrachi, alongside a team of scientists under the mentorship of Permutt, published their findings in Nature (journal) regarding a mutation that lead to Wolfram syndrome (WFS). The study used linkage analysis in order to characterize WFS amongst 3 Japanese and 2 European families. D4S500 and D4S431 were located on the same yeast artificial chromosome (YAC) as previously identified in the Stanford Chromosome 4 YAC mapping project. Using human P1 and bacterial artificial chromosome (BAC) clones, a contiguous fragment was constructed encompassing the WFS critical region to appropriately map the region. A method called exon trapping, which is now replaced by computational-based cDNA sequencing, was used to determine potential exons in unknown DNA fragments through insertion into a splicing vector. Large-scale genomic sequencing enabled for characterization of 180 kb, which was eventually narrowed to an ORF of 2672 nt (890 aa) via PCR amplification and corresponding northern blotting with a poly A+ RNA probe. This gene was named WFS1, and was compared in patients with and without Wolfram syndrome. The primary conclusion was a homozygous frameshift mutation that lead to the creation of an elongated 937 aa product, among a host of other polymorphisms. WFS1, was as Bernal-Mizrachi helped determine, crucial for appropriate pancreatic β cell function, and accumulation of mutated WFS1 isoforms induced early-onset insulin deficient diabetes mellitus.

Akt Pathway Signaling and Effect on β cell Mass

In 2001, Bernal-Mizrachi turned his attention towards specific signaling pathways and their effects on β cell function, publishing his work in the Journal of Clinical Investigation. His goal was to study the in-vivo effects of phosphoinositide 3-kinase (PI3K) Akt/PKB pathway. Akt is a serine-threonine kinase that phosphorylates other targets that lead to cell growth and proliferation, and is activated by PI3K. Bernal-Mizrachi used a transgenic mouse model, in which a constitutively active Ak1 gene, which lacked a PH domain that otherwise mediates interaction with PI3K, was fused with a hemoglutinin tag (HA) for checking expression levels. An pre-prepared RIP/β-globin expression vector was created, and the AK1 construct was inserted at an EcoRI restriction site. This chimeric gene was then injected into embryonic stem cells in fertilized mice eggs.

To approximate β cell Mass in control and transgenic mice, the pancreas was excised and immunohistochemistry detected insulin-positive cells in tissue sections. A guinea pig anti-human insulin was hybridized to insulin, and 3-amino-9-ethyl carbazole (AEC) was the conjugated chromophore that could be detected via fluorescence microscopy. Image analyze software from BioQuant allowed for a determination of the ratio of β cell area to total area, which was multiplied by pancreatic gross weight to quantify β cell mass. Glucose levels were also measured after injection following a 6-hour fast, and a rat insulin ELISA kit. Results indicated that β cell mass in transgenic mice increased eight to nine-fold, suggesting Akt overexpression drives β cell neogenesis pathways. Additionally, the fasting state provoked a 1.7 fold increase in plasma insulin, leading to enhanced glucose tolerance.

Other Significant Discoveries (2000 - Present)

Building upon his prior work with the Akt pathway, Bernal-Mizrachi took a closer look at the dysregulation in insulin secretion that resulted from reduced pathway activation. The negative control, or kinase-dead Akt1, was inserted into a RIP-I/β-globin expression vector, and then injected into fertilized mice eggs. This transgene was confirmed to be expressed in β cells using immunofluorescence staining and Western blot analysis with anti-HA (hemagglutinin) and anti-insulin Ab. Using an in vitro kinase assay, the phosphorylation of two proteins (S6K and Foxo1) could be observed, and kinase dead Akt lysates as expected, had significantly lower rates of phosphorylation. His key finding was in the abnormal patterns of insulin secretion resulting from Akt dysregulation. A series of islet perifusion experiments were performed, where groups of islets were bathed in various glucose concentrations (2 mM basal, 20 mM). Upon this higher glucose concentration, reductions of insulin secretion were observed, even when treated with a Ca(2+) ionophore ionomycin that increases Ca(2+) intracellular concentration. Therefore, the role of the Akt pathway was concluded to be related to the exocytosis mechanism of insulin.

Primary Honors and Awards

Bernal-Mizrachi is a member of the American Society for Clinical Investigation (ASCI) since 2010, and won a Junior Faculty and Career Development Award from the American Diabetes Association. He also is a recipient of multiple NIH grants as well as the Veterans Affairs Biomedical Laboratory R&D Senior Clinician Scientist Investigator Award in 2022.

Furthermore, he is also board-certified in internal medicine and endocrinology [2]. Bernal-Mizrachi, in addition to research, works as a physician at the Miami Veterans Administration Hospital.

Current Lab Focus

After spending decades elucidating the PI3K/Akt pathway manner of induction of β cell proliferation and mass, Bernal-Mizrachi has expanded his focus on the broader impacts of type 2 diabetes treatment. He is also interested in how to mitigate cell death with β-cell transplantation and has worked to develop transgenic mice models for transplantation. The ultimate goal of his lab, which complements his clinical practice, is to further understanding of pancreatic cell specialization and probe novel remedies (pharmacologic and transplant-based) to help eliminate the need for standard medicine/insulin-based therapies.

References

[2] https://med.miami.edu/faculty/ernesto-bernalmizrachi-md#:~:text=Bernal%2DMizrachi%20has%20received%20many,for%20Clinical%20Investigation%20(ASCI).

[3] https://www.jci.org/articles/view/13785 ~ Akt transgenic mice

[4] https://www.nature.com/articles/ng1098_143 ~ 1998 Wolfram Gene

[5] https://www.jci.org/articles/view/20016 ~ Akt mutant --> exocytosis failure

[6] https://umiamihealth.org/en/sylvester-comprehensive-cancer-center/research/labs/bernal-mizrachi-lab ~ Current Lab Work