This report introduces the Nile rat as a novel model of Metabolic Syndrome associated with diet-induced diabetes in a readily managed lab rodent of intermediate size. The diabetes was age, sex, and diet dependent, and appeared related to stressful gene-environment interactions that affected energy utilization. It is unique in its high percentage of involvement (close to 100% in males, somewhat less in females), and in the fact that several aspects of the disease examined in this report (early abdominal obesity, hyperinsulinemia, elevated blood glucose and triglyceride plus hypertension, as well as unpublished data on depressed HDL) mimic Metabolic Syndrome and type 2 diabetes in humans [9, 12–15].
Diet and Growth considerations
Several aspects of diabetes in Nile rats are noteworthy from a nutritional point of view, particularly the disposition of energy for growth or storage as adipose. Specifically, males fed a chow diet (Lab Diet, #5020, 3.57 kcal/g) grew faster than females (as expected), and they developed diabetes more readily than females, presumably based on sex hormone differences. Even among individual males, those that grew faster, reminiscent of human infants with a rapid postnatal weight gain [16–19], developed diabetes sooner (early-onset groups).
One might assume that more rapid weight gain represented greater food consumption leading to adipose accumulation and diabetes. However, only for a brief 2 wk period in the youngest weanlings, age 5 to 7 wks (Expt 6), could we detect early-onset diabetes linked to greater food intake (20%) and greater adiposity (37%). Thus, linear growth was faster (body length), relative muscle mass less (carcass % bd wt) and fat depots greater (adipose % bd wt) for early-onset rats. But even there, greater growth efficiency (33% better) had a greater impact than the increased calorie intake. In a second experiment where food intake was tracked for 32 wks beginning at 8 wks of age (Expt 3), calories consumed did not differ as a function of diabetes onset, but better growth efficiency (100%) still pertained for the early-onset group. Expt 3 also teaches that growth efficiency, as a causal link for diabetes onset, only applied during rapid growth, as the relationship between calorie intake and body weight gain disappears once growth ceases. When growth is complete and diabetes established, calorie intake and efficiency measures of energy utilization for growth would no longer pertain.
In late stages of the disease in older rats (Table 2, 11 mo), blood glucose and plasma lipids were severely elevated, and the rats developed insulin-dependent diabetes with adipose wasting, especially in the perirenal fat pad, even as calorie intake doubled. This combination of wasting adipose plus elevated plasma triglycerides in advancing diabetes, mimics the defect of free fatty acid recycling seen in hyperinsulinemic humans with insulin resistance and diabetes , confirming the comparable metabolic profile in these Nile rats.
Thus, rats that used calories more efficiently to (atypically) accelerate linear growth and early weight gain, including adipose accumulation relative to muscle mass, were more prone to insulin resistance and diabetes later on. In addition, a shift to fat catabolism (especially perirenal fat) in the later stages of diabetes may have complicated our estimates of adiposity in cross-sectional studies, since adipose tissue may increase initially then begin to decline, depending on the rat age and stage of diabetes. It is important to note that, in general, the diabetes did not appear to depend on hyperphagia or generalized obesity per se, as total body fat never exceeded 10 to 15% of body weight and, in most experiments, did not correlate with blood glucose. This differs substantially from most other mouse or rat models of diabetes.
Wild Nile rats
The relationship between rapid growth and diabetes is supported by the observation that Nile rats in the wild, where food and calories are less available, grow less rapidly than captive rats and do not develop diabetes. In fact, body weight of wild Nile rats, even in the rainy season when food is more abundant, was about 35% less in males (ie. 85 g final wt) and 40% less in females (65 g final wt) than comparable weights of captive-fed rats . During the dry season both male and female wild rats weighed about 60% less than captive-fed rats. Nonetheless, wild rats reach their reduced adult weight in approximately the same amount of time (5 mo) as captive rats. These differences also reflect the fact that their grasses-and-insects diet in the wild has a caloric density close to 2.0 kcal/g , compared to the 3.2-4.5 kcal/g in our lab diets. Thus, if food is curtailed by natural environmental conditions linked to the dry season  or by food restriction in the laboratory setting, diabetes risk is reduced.
To this point, two of the present experiments demonstrate that dietary factors influenced the onset and degree of diabetes observed, similar to the situation for type 2 diabetes in humans  and sand rats [3, 5, 22, 23]. On the one hand, reducing calorie intake to 75% ad libitum affected weight gain only slightly (n.s.), but limited diabetes, protecting against fatal disease much as modest weight loss does in humans with type 2 diabetes . Sand rats reportedly even reverse their diabetes if food intake is restricted 50% before beta-cell failure occurs [3, 25]. In the second instance a high-energy, Western-type diet (4.5 kcal/g), which would potentiate fatty acid oxidation and fat storage, significantly elevated blood glucose and rendered Nile rats severely glucose-intolerant relative to a low-fat, high-fiber Mediterrean-type diet (3.5 kcal/g). Interestingly, despite these differences in dietary energy density and composition, rats compensated with the result that their total energy intake and body fat pools did not differ for the 24 wk comparison. Furthermore, even though the random blood glucose at study end was highest for the low-fat (high-CHO) group, that group also had the lowest fasting glucose (Table 5). This suggests that the low-fat diet still allowed for reserve insulin secretion to reduce an elevated random glucose during the 16 h fast. Thus, the rate and composition of energy processed by the Nile rat influences type 2 diabetes onset without necessarily revealing outward signs on body weight and adiposity. From these observations it is not surprising that food restriction and weight loss are the first method of choice for alleviating diabetes in humans afflicted with type 2 diabetes .
"Thrifty genes" and insulin resistance
The response to restricted calorie intake and enhanced calorie utilization for growth in Nile rats with early-onset diabetes is also reminiscent of the sand rat, where evidence for the "thrifty genes" theory was presented to explain the prevalence of diabetes in a susceptible strain, compared to a more resistant one [3, 12, 26, 27]. In that situation diabetes-prone sand rats experienced 33% greater feed efficiency and became insulin resistant, eventually storing more energy as body fat per calorie consumed than the resistant strain. Unfortunately, no data were presented for carcass or muscle mass representing somatic growth rate. In the same manner, both older and younger cohorts of Nile rats with early-onset diabetes (Expts 3 and 6) were more energy efficient than the late-onset groups of the same age; and those in Expt 6 also accumulated more fat early, indicating that the early-onset group (and presumably expression of their Metabolic Syndrome) was related to genetic control of energy utilization, particularly in young rats.
These data suggest that genetically controlled growth rate is modulated by diet composition, including dietary calorie density  and availability, possibly including the specific macronutrient composition of the diet itself. For example, high-fat, low-fiber diets are notorious for inducing diabetes in susceptible models . Once growth ceased and energy shifted from linear skeletal growth and muscle expansion to maintenance/storage, it would appear that insulin resistance (initiated during rapid growth) led to sustained hyperglycemia . Our late-onset rats simply grew slower, so they delayed their destiny with diabetes because they required more calories for less growth (less efficient) early on, presumably reducing calories available for storage. In sand rats, which are ecologically and physiologically similar to Nile rats [4, 23, 25], insulin resistance precedes hyperglycemia, similar to humans and Nile rats . One current theory suggests that excessive fatty acid oxidation in muscle mitochondria leads to ROS-induced damage that initiates insulin resistance as a means to protect muscle from the burden of additional energy disposal [29, 30]. Another theory suggests that excessive uptake of free fatty acids by tissues, including inflammation and death of pancreatic beta-cells, leads to insulin resistance and fat accumulation in liver and muscle . To date the role of inflammation has not been explored as a factor contributing to the Metabolic Syndrome of Nile rats.
Similar to early "growth stress" in Nile rats, epidemiological studies have noted that small-for-term infants growing faster at 7 years of age than infants with normal birth weight were more likely to develop type 2 diabetes as adults . It is thought that in utero nutrition is a key dynamic [33, 34], but nothing is known about the specifics of the nutrients involved or the character of postnatal nutrition, other than high energy intake being a risk factor affecting the incidence and onset of their diabetes . It will be important to define the maternal metabolism and nutrient intake of the Nile rat during pregnancy, as well as details relative to postnatal diet composition and behaviors around food intake that impact the growth rate of their pups, particularly males.
In any event the Nile rat model, with a focus on growth and food (energy) utilization during rapid growth between 3-10 wks of age, provides an opportunity to identify diet-gene interactions underlying insulin resistance and diabetes. With this paradigm, it should be possible to intervene with diet or drug to establish relative efficacies for prevention of overt, diet-induced insulin resistance and diabetes in experiments of 4-7 weeks, which is unique among animal models fully expressing this disease from natural causes.
Animal models are useful for studying the pathophysiology of human disease and allow for therapeutic intervention with an abbreviated time span relative to human experiments. The model should mimic the human disease, or at least present its major symptoms for study. So far no animal model mirrors all characteristics of the Metabolic Syndrome and non-insulin-dependent (type 2) diabetes mellitus . Many mouse and rat models have idiosyncratic similarities to certain aspects of the human condition, often attributed to a specific gene mutation. For example, the ob/ob [obese], db/db [diabetes] and the Zucker diabetic fatty rat (ZDF) have mutations either in the leptin gene (ob/ob) or in the leptin receptor (db/db) which causes over-eating obesity, unlike the natural history of the disease in humans where leptin is seldom implicated [37, 38]. Gene analysis through backcross breeding of diabetes-resistant and diabetes-prone sand rats suggests that a single major gene may control the transition from normo- to hyperglycaemia in that model . Still another sand rat report labeled larger male rats as "obese" (with <6% of total body weight as fat), and suggested their "obesity" was the reason for their insulin resistance and diabetes. However, differential growth measures distinguishing between lean body growth (lean mass per se) and BMI (adiposity) were lacking, so the question remains whether these desert rodents develop diabetes due to accelerated growth rates or overabundant fat accumulation, or both . The C57BL/6 mouse is more susceptible to diet-induced obesity (DIO) and diabetes than other mouse strains , but its diabetes is not as progressive or severe as that in either the Nile rat or sand rat, and does not present all sequelae present in the human disease. This is true even though the DIO mice often double their weight from added adipose. Plus, clinical aspects of diabetes in DIO mice are subtle and of minimal clinical consequence, taking 6-12 mo to develop. Extreme overall obesity is not characteristic of diabetes in Nile rats or sand rats, evidenced by the 10 to 15% upper limit in body fat observed herein, and the 2-6% range in male or female sand rats .
The utility of the Nile rat model should be emphasized from several aspects. First, it appears to be a polygenic wild-type rodent (outbred) that expresses all aspects of the Metabolic Syndrome measured to date, including abdominal fat accumulation (at least in young rats), hyperinsulinemia, hyperglycemia, hypertriglyceridemia (with depressed HDL, unpublished data), microalbuminuria (data not shown), and hypertension on the way to terminal type 2 diabetes. Second, as in the sand rat , the diabetes has 5 clearly marked stages leading to ketosis and death, which are analogous to human type 2 diabetes. Pathology of the liver, kidney, and pancreatic beta cells appear to be similar to human disease  and invite investigation of issues still outstanding around type 2 diabetes in humans, eg. mechanisms and interventions to improve insulin secretion or kidney function associated with normal progression of the diet-induced disease. Third, the incidence of diabetes is high in this captive-bred model, slightly greater than that reported for out-bred sand rats . Incidence in the sand rat has been enhanced further by selective inbreeding in an Israeli colony . Almost all captive male Nile rats fed chow become diabetic by 1 year, some as early as 6-10 wks. In weanling sand rats diabetes developed in a susceptible inbred strain within 2 wks on a challenge diet (only 2.9 kcal/g vs. our Nile rat challenges of 3.57 kcal/g in chow or 3.5 kcal/g and 4.5 kcal/g in our purified diets) followed by ketosis and death in 10 wks . By contrast, to date in our older Nile rat cohorts, even our higher density diet produced a less acute, more prolonged, but all-inclusive disease in males. Female Nile rats delay diabetes onset, with approximately 75% eventually developing the disease by 1-1.5 yr. Similarly, severity and incidence of diabetes in captive female sand rats was slightly less than males at 16 wk of age when fed a chow diet in an outbred colony . Many data suggest that type 2 diabetes may be more common in men than women, at least premenopausal women [32, 42, 43].
Certain physiological correlates of the diabetes in Nile rats are noteworthy. First, blood glucose was highly correlated with hepatomegaly, which reflected hepatic steatosis similar to humans with type 2 diabetes . Second, kidney enlargement with polyuria, and eventual renal failure ending in glomerular and interstitial nephritis and sclerosis with reduced kidney size were associated with polydypsia, polyuria, and ketosis at the endstage of the disease linked to elevated glucose (unpublished data). Diabetes progression also was associated with elevated blood pressure, which is a frequent morbid association in humans with diabetes [45, 46]. These observations, coupled with the hyperlipemia that develops as glucose rises, suggests that abnormal energy metabolism based on liver handling of insulin and calories leading to insulin resistance are critical aspects of the disease in Nile rats, even as they are in humans with the type 2 diabetes and Metabolic Syndrome .
Random versus fasting glucose
One practical issue examined was whether random or fasting glucose is the better measure of this diabetes. While both measures demonstrate disease progress, the random value is easier to execute, as it does not require prolonged fasting. Because Nile rats are diurnal, they eat predominantly during the day, so a random glucose value is never far removed from eating, which can be considered a continuous stimulus for insulin secretion during daytime . Furthermore, first signs of glucose intolerance (insulin resistance) can be detected easier with excursions in random blood glucose than with fasting glucose. A 16 h fasting glucose value, on the other hand, provides the opportunity to determine whether enough insulin can still be generated to clear circulating glucose, thereby demonstrating the reserve capacity of beta-cell function in the host. When insulin secretion is waning in Nile rats, fasting glucose remains elevated, even as it does with fasting in advanced type 2 diabetes in humans [14, 24, 48].
Thus, in this model energy (glucose, triglycerides) is not removed from systemic circulation effectively despite elevated insulin (hyperinsulinemia reflecting insulin resistance), which leads to a blood glucose increase that eventually leads to beta-cell failure with decreased insulin production [47, 49, 50]. The cutoff points for blood glucose differ for fasting or random glucose, but they were highly related. Random glucose above 150 mg/dl was a good predictor of diabetes from the current data, while fasting glucose >110 mg/dl seems reasonable for diabetes onset. Random glucose commonly rises to 300-500 mg/dl, often over 600 mg/dl, in Nile rats with advanced diabetes and insulin depletion, whereas fasting glucose seldom exceeds 400 mg/dl, even in the worst case of diabetes. Slower progressing diabetes in older Nile rats also was associated with a much greater hyperglycemia and hyperlipemia, with elevated VLDL-TG (up to10-fold), total cholesterol (5-fold) and HDL depressed to 1/5 normal (unpublished data). By contrast, triglycerides and cholesterol reportedly rise only minimally in sand rats .