**3. The role of obesity in the development of type 2 diabetes**

The decrease in insulin action indicates insulin resistance, the main hallmark of T2DM [38]. Several factors play a role in the development of insulin resistance, with obesity probably being the most important one [39]. During the last three decades, obesity has reached an epidemic stage in all age groups [40]. Increased intake of energy-dense foods containing a high percentage of fat and carbohydrates, combined with a lack of physical activity, leads to a net positive energy balance and represents the primary cause of obesity. The first stages of obesity are hypertrophy and hyperplasia, where adipocytes try to meet the demand to store excessive energy. When levels of free fatty acids and triglycerides exceed the metabolic capacity of adipose tissue, they accumulate as ectopic fat in non-adipose tissue such as the liver, pancreas, skeletal muscles, and heart [41]. The regional distribution of adipose tissue is particularly important for the development of disorders of glucose and lipid metabolism [42, 43]. The fat infiltration in the liver and pancreas in the absence of excess alcohol intake is termed non-alcoholic fatty liver disease (NAFLD) and non-alcoholic fatty pancreas disease (NAFPD), respectively [44]. Due to fat accumulation in hepatocytes, NAFLD leads to inflammation, fibrosis, cirrhosis, and liver cancer and critically affects insulin sensitivity in the liver [44–46]. NAFPD plays a similar role in the dysfunction of the endocrine and exocrine pancreas leading to exacerbation of acute pancreatitis and increasing the susceptibility to pancreatic cancer [46–50]. Although the concept of the NAFPD was introduced only a few years ago [51], the correlation between fat infiltration and pancreas was first described almost a century ago on

obese cadavers having larger pancreata compared to non-obese cadavers [52, 53]. Importantly, over the last decade, it has been demonstrated convincingly that most of the morphological, functional, and clinical features of T2DM are reversible with sufficient weight loss and that these positive changes depend on the level of hepatic and pancreatic fat reduction [54–56].

Although insulin resistance is one of the main features of T2DM, it is not sufficient for the development of T2DM. Despite increasing insulin resistance, present long before the onset of diabetes, individuals with preserved beta cell capacity can stay normoglycemic for several years due to compensatory insulin hypersecretion. Furthermore, a certain proportion of individuals with insulin resistance never develop T2DM. The idea has been put forward that there may be a personal fat threshold or an individual level of susceptibility to developing T2DM at a given body mass index, with the main mechanisms behind this attractive hypothesis being varying degrees of fat accumulation in the liver and the pancreas, together with varying individual responses to this accumulation [57]. An obese subject with insulin resistance can secrete 2–5 times more insulin compared to lean nondiabetic individuals in response to a glycemic load, but when the adaptive capacity of the beta cells fails, T2DM occurs [58]. Several studies have shown that the first phase of insulin secretion is primarily affected, resulting in impaired glucose tolerance. During the progression of T2DM, the second phase of insulin secretion is further lost. Post-translational defects in insulin synthesis also occur, resulting in increased proinsulin secretion, and by the time the diagnosis of T2DM is made, the beta cell function is already typically reduced by 80% [59]. This beta cell failure is responsible for transitioning from an insulin-resistant compensated state to overt T2DM and remains to be elucidated in details. It probably involves an initially inadequate beta cell mass and an insufficiently increased response of the existing beta cells to increased insulin demand [60]. Noteworthy, the susceptibility of beta cells themselves to developing insulin resistance may play an important pathophysiological role [61]. Individuals with T2DM can display changes in beta cell mass, either due to a decrease in beta cell proliferation or/and an increase in cell apoptosis [62]. In T2DM patients, the apoptosis rate is increased severalfold compared to normoglycemic individuals [63], a phenomenon confirmed by ultrastructural analysis using TEM [64]. Furthermore, during T2DM gradual dedifferentiation of beta cells occurs in animal models, while the role of dedifferentiation during the development of human T2DM is controversial and less well-studied [63, 65–68]. Chronic exposure of islets to elevated levels of glucose, fatty acids, and amino acids results in ER stress due to increased insulin synthesis and secretion [69]. The protein folding capacity of ER becomes exceeded, which leads to the activation of the so-called unfolded protein response (UPR) and consequently to the inhibition of protein translation [70, 71]. Gluco- and lipotoxicity cause a vicious cycle of continuous deterioration of the glucometabolic state and eventually impair insulin secretion and increase apoptosis. In T2DM, not only the islet cells are affected, but also the pancreatic acinar cell viability and growth decrease, leading to increased apoptosis and replacement by fat [72, 73]. Importantly, intracellular fat in acinar cells may affect beta cell function in a paracrine manner through the release of adipokines. It is also likely to play a role in pancreatic carcinogenesis, seems to be associated with changes in innervation, and may initiate an acinar-to-adipocyte trans-differentiation [56].
