Editorial
Diabetes and obesity have emerged as major public health burdens and raise the risk of neurological dysfunction, leading to cognitive impairment in patients1. As a result of the alarming rate of increase in these diseases, research efforts to combat these health challenges are growing. This methods collection brings together a wide range of techniques that are used to study diabetes and obesity, along with the associated cognitive impairment, in a variety of model systems.
The zebrafish is an attractive model to study diabetes because the expression and function of the genes that are involved in pancreas development are highly conserved in this organism when compared to mammalian systems2,3,4,5,6. Diabetes mellitus and hyperglycemia can be induced in zebrafish using invasive and non-invasive mechanisms and by developing mutant strains7,8,9,10,11. McCarthy et al. introduce a novel, non-invasive method to induce hyperglycemia in zebrafish using an alternate immersion protocol12. In this method, adult zebrafish are placed in a sugar solution for 24 h and then in water for 24 h to mimic the rise and fall of blood sugar in diabetes. They demonstrate that this process not only successfully induces hyperglycemia but also leads to the complications related to altered glucose homeostasis. Once established, this method could be successfully used to mimic the hyperglycemic condition in adult zebrafish.
Uncontrolled hyperglycemia can lead to an array of complications affecting every organ in the body. Changes in memory and/or cognitive impairment are among the concerning effects of uncontrolled hyperglycemia in patients with elevated blood glucose. As zebrafish have successfully been used in studies testing the effects of altered glucose homeostasis13, this organism should now be used in behavioral studies as well. In their second article, Rowe et al. demonstrate a method to use zebrafish in behavioral studies14 using a modified three-chambered apparatus designed to monitor social interactions in the mouse15. The zebrafish demonstrate acquired memory in about 8 weeks of training. However, the hyperglycemic fish demonstrate cognitive deficits after 4 weeks of treatment to induce hyperglycemia using an alternate immersion protocol12. This method provides a way to use zebrafish in behavioral studies that will be useful not only in the field of diabetes but also in other fields related to age-related, genetic, and other neurodegenerative diseases that lead to cognitive impairment.
Along with the ongoing research on the disease etiology and its effects, the field needs novel molecular avenues to generate functional β-cells for therapeutic interventions. However, the successful production of β-cells, either by in vitro culture or by the stimulation of in vivo regeneration, has been limited. Memon and Abdelalim describe a method of differentiating human pluripotent stem cells (hPSCs) into PDX1- (pancreatic and duodenal homeobox 1) and NKX6.1 (NKX6 homeobox 1)-expressing pancreatic progenitor cells16. The expression of these two transcription factors in progenitors reduces the occurrence of non-functional endocrine cells in cultures and results in functional, insulin-producing, glucose-responsive β-cells17. The scalable generation of PDX1 and NKX6.1 co-expressing progenitors has the potential to generate a functional pool of insulin-secreting β-cells for transplantation therapy as well as for disease modeling.
Obesity is a complex metabolic disorder caused by abnormal adipogenesis. Due to the increased need for human adipocytes to model the disease conditions and the limitations of collecting samples from human donors, the in vitro generation of adipocytes is in great demand. Aghadi et al. describe a protocol to produce mesenchymal stem cells (MSCs) from embryoid bodies derived from induced pluripotent stem cells (iPSCs) using all-trans retinoic acid18. The resulting MSCs show rapid proliferation and successful differentiation into adipocytes. They also establish a method to increase the differentiated adipocyte pool by sorting the cells tagged with Nile red, a lipophilic dye18. Taken together, they successfully produced cultures enriched in a pure population of mature adipocytes that will help researchers to study adipocyte-associated metabolic disorders with ease.
There are two major problems in the field of obesity research: the inability of cultured cell lines such as 3T3-L1 to reflect the cellular heterogeneity of the adipose depot19,20, and the low yield of preadipocytes from the tissue collected from adult mice21,22. The cells collected from adult mice can reflect the cellular heterogeneity but contain mostly fully matured adipocytes and fewer preadipocytes21,22. Saez et al. answer both problems by describing a method for isolating white and brown preadipocytes from newborn mice23. These cells show high proliferative capacity and differentiation potential into mature adipocytes compared to the preadipocytes collected from adult mice. Their article is an effective resource to model the complexity of obesity using white and brown preadipocytes that can be differentiated into fully mature adipocytes.
Weight management is an important clinical issue. Long-term lifestyle changes, anti-obesity drugs, and exercise regimes all play important parts in this process. However, the loss of motivation due to the challenges in maintaining long-term lifestyle modifications, the expense of anti-obesity drugs, and the increased incidence of eating disorders have led the scientific community to explore natural remedies24. Pérez Gutiérrez and Arrioja explore the potential of two plants, Syzygium aromaticum (clove) and Cuminum cyminum (cumin)25, as natural resources of anti-obesity compounds. The active phytochemical contents are extracted using ultrasound in an ethanol:water (50:50) solution and administered to C57BL6/J mice fed a high-fat diet. This represents an obesity model. The outcomes include improvements in various obesity parameters such as the regulation of lipid profiles (cholesterol and triglycerides) and reductions in food intake, weight gain, adipose tissue weight, and liver weight. This proposed method takes 5 weeks compared to the traditional 16 week regime and has the potential to test and identify new phytochemicals for their possible use in weight management.
This methods collection provides novel ways to use zebrafish in behavioral studies, to induce a hyperglycemic condition in this model organism, and to generate progenitor cells that lead to the production of insulin-producing, glucose-responsive β-cells. The collection also focuses on obesity and introduces methods to successfully produce adipocytes from iPSCs, to collect preadipocytes from newborn mice, and to isolate phytochemicals to test their potential in treating obesity. Taken together, this collection targets a broad group of researchers in the field of diabetes and obesity and provides a novel toolkit to illuminate previously unexplored areas in diabetes and obesity research.
Disclosures
The author has nothing to disclose.
Acknowledgments
I thank the authors for enriching the fields of diabetes and obesity with the novel methodologies and the Journal of Visualized Experiments for helping in this process.
References
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- Argenton, F., Zecchin, E., Bortolussi, M. Early appearance of pancreatic hormone-expressing cells in the zebrafish embryo. Mechanisms of Development. 87 (1-2), 217-221 (1999).
- Biemar, F., et al. Pancreas development in zebrafish: Early dispersed appearance of endocrine hormone expressing cells and their convergence to form the definitive islet. Developmental Biology. 230 (2), 189-203 (2001).
- Lin, J. W., et al. Differential requirement for ptf1a in endocrine and exocrine lineages of developing zebrafish pancreas. Developmental Biology. 270 (2), 474-486 (2004).
- Yee, N. S., Yusuff, S., Pack, M. Zebrafish pdx1 morphant displays defects in pancreas development and digestive organ chirality, and potentially identifies a multipotent pancreas progenitor cell. Genesis. 30 (3), 137-140 (2001).
- Zecchin, E., et al. Evolutionary conserved role of ptf1a in the specification of exocrine pancreatic fates. Developmental Biology. 268 (1), 174-184 (2004).
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- Agardh, E., Bruun, A., Agardh, C. D. Retinal glial cell immunoreactivity and neuronal cell changes in rats with STZ-induced diabetes. Current Eye Research. 23 (4), 276-284 (2001).
- Carmo, A., Cunha-Vaz, J. G., Carvalho, A. P., Lopes, M. C. Nitric oxide synthase activity in retinas from non-insulin-dependent diabetic Goto-Kakizaki rats: Correlation with blood-retinal barrier permeability. Nitric Oxide. 4 (6), 590-596 (2000).
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- Sharchil, C., et al. Zebrafish: A model to study and understand the diabetic nephropathy and other microvascular complications of type 2 diabetes mellitus. Veterinary Sciences. 9 (7), 312-327 (2022).
- Rowe, C. J., Crowley-Perry, M., McCarthy, E., Davidson, T. L., Connaughton, V. P. The three-chamber choice behavioral task using zebrafish as a model system. Journal of Visualized Experiments. (170), e61934 (2021).
- Crawley, J. N., et al. Automated apparatus for quantitation of social approach behaviors in mice. Genes, Brain and Behavior. 3 (5), 303-314 (2004).
- Memon, B., Abdelalim, E. M. Differentiation of human pluripotent stem cells into pancreatic beta-cell precursors in a 2D culture system. Journal of Visualized Experiments. (178), e63298 (2021).
- Memon, B., Abdelalim, E. M. Stem cell therapy for diabetes: Beta cells versus pancreatic progenitors. Cells. 9 (2), 283-302 (2020).
- Aghadi, M., Karam, M., Abdelalim, E. M. Robust differentiation of human iPSCs into a pure population of adipocytes to study adipocyte-associated disorders. Journal of Visualized Experiments. (180), e63311 (2022).
- Gregoire, F. M., Smas, C. M., Sul, H. S. Understanding adipocyte differentiation. Physiological Reviews. 78 (3), 783-809 (1998).
- Schoettl, T., Fischer, I. P., Ussar, S. Heterogeneity of adipose tissue in development and metabolic function. Journal of Experimental Biology. 221, (2018).
- Arner, P., et al. Dynamics of fat cell turnover in humans. Nature. 453 (7196), 783-787 (2008).
- Wang, Q. A., Tao, C., Gupta, R. K., Scherer, P. E. Tracking adipogenesis during white adipose tissue development, expansion and regeneration. Nature Medicine. 19 (10), 1338-1344 (2013).
- Galmozzi, A., Kok, B. P., Saez, E. Isolation and differentiation of primary white and brown preadipocytes from newborn mice. Journal of Visualized Experiments. (167), e62005 (2021).
- Sun, N. N., Wu, T. Y., Chau, C. F. Natural dietary and herbal products in anti-obesity treatment. Molecules. 21 (10), 1351-1365 (2016).
- Pérez Gutiérrez, R. M., Arrioja, M. W. Rapid model to evaluate the anti-obesity potential of a combination of Syzygium aromaticum (clove) and Cuminum cyminum (cumin) on C57BL6/j mice fed high-fat diet. Journal of Visualized Experiments. (173), e62087 (2021).
