In the light of a prior study conducted in sand arts [11
], the major aim of the present experiments was to investigate the potential benefit of intermittent fasting in STZ-induced diabetic rats. Parallel experiments were here conducted in control animals.
As expected, the glycemia (or plasma D-glucose concentration) and the total AUC during an IPGTT were much higher in STZ rats than in control animals. Even the incremental AUC during the IPGTT was higher (P
< 0.006) in STZ rats (608 ± 105
= 52) than in control animals (287 ± 29
= 48) despite a comparable initial increment in glycemia 30
min after the injection of D-glucose. Also, as expected, the plasma insulin concentration, the insulinogenic index, the secretion of insulin by isolated islets, their insulin content, the relative volume occupied by the β
-cells in serial sections of the whole pancreas and the individual islet area were much lower in STZ rats than in control animals. Incidentally, a positive secretory response to D-glucose was still observed in isolated pancreatic islets from STZ rats, this coinciding with a progressive decrease in their final insulin content after incubation at increasing concentrations of D-glucose. Such a decrease was not observed, however, in pancreatic islets from control animals. Last, the HOMA for insulin resistance and percentage of apoptotic β
-cells were also significantly higher in STZ rats than in control animals.
In terms of morphological findings, there was, as a rule, little to distinguish between NF, IF, and CR control animals. At the most, there was a trend (P < 0.02) towards a higher individual β-cell area in IF than in NF control rats. Moreover, the pancreatic wet weight and, hence, total β-cell mass appeared lower (P < 0.04 or less) in IF control rats than in NF control rats. A comparable situation (P < 0.02) prevailed when comparing the pancreatic wet weight in IF STZ rats and NF STZ rats.
In the STZ rats, the major other changes attributable to differences in feeding schedule concerned, in terms of morphological findings, the relative and absolute values for β-cell mass, the individual β-cell and islet area and the percentage of apoptotic β-cells. In the IF STZ rats, the relative and absolute values for β-cell mass, as well as the individual β-cell area and islet area, were all higher than in NF STZ rats, whilst the percentage of apoptotic cells appeared lower in IF than NF STZ rats. In the CR STZ rats, comparable findings were restricted to the individual islet area and percentage of apoptotic cells.
The much higher percentage of apoptotic β
-cells in STZ rats, as compared to control animals, is likely attributable to two major factors. First, the β
-cell cytotoxic effect of STZ should not be ignored. According to Morimoto et al. [22
], apoptosis of β
-cells is already detected 6 hours after the injection of STZ, even before the onset of hyperglycemia. In this respect, it should be kept in mind that 3 to 14 days after STZ administration, infiltration of the islets by mononuclear cells takes place, eventually resulting in the removal of apoptotic cells by unspecific macrophages [23
]. Hence, the present data may well underestimate the apoptosis index otherwise prevailing during the first days after STZ administration. A second factor consists of the hyperglycemia resulting from STZ administration, since in vitro
exposure of pancreatic islets to high concentrations of glucose also induces β
-cell apoptosis [24
The latter process may account, in part at least, for the apparent differences between IF and CR STZ rats, in terms of both relative β-cell mass and individual β-cell area. Thus, according to the data listed in Tables and , the glycemia (or plasma D-glucose concentration) after overnight starvation, expressed relative to the mean corresponding values found in NF STZ rats averaged in the IF STZ rats 64.9 ± 5.7% (n = 21), as distinct (P < 0.01) from 86.6 ± 5.6% (n = 23) in the CR STZ rats. The mean values for both the total and incremental AUC during the IPGTT were also higher in CR STZ rats than in IF STZ rats (). Such differences in glucose homeostasis coincided with higher mean values for both the relative volume of β-cells and their individual area in IF STZ rats as distinct from CR STZ rats (Tables and ). Thus, the mean relative value of β-cells was almost twice higher in IF than CR STZ rats, whilst the mean individual β-cell area represented in the IF STZ rats 125.4 ± 6.0% (n = 20; P < 0.002) of that found in CR STZ rats (100.0 ± 4.0%; n = 20). Despite the vastly different magnitude of the IF/CR ratio for these two variables, the difference between CR and IF STZ rats remained highly significant (P < 0.005) when pooling together the results recorded for each of these variables.
The individual β
-cell area was also always significantly higher in STZ rats than in control animals exposed to the same dietary schedule (). These converging findings concerning differences in individual β
-cell area as a function of glucose tolerance are reminiscent of the hypertrophy of β
-cells found either in vitro
after exposure to a high concentration of D-glucose [25
] or in vivo
in rats which became hyperglycemic after partial pancreatectomy [26
] and currently ascribed to a compensatory mechanism in residual β
-cells no more susceptible to undergo mitosis [27
The latter consideration is not meant to deny that in addition to β
-cell hypertrophy, an increase in β
-cell number, possibly attributable to transdifferentiation of glucagon-producing to insulin-producing cells [28
], may participate in the difference in relative or total β
-cell mass between NF and IF STZ rats, as also suggested by the total cell numbers listed in .
A beneficial effect of intermittent fasting from 5
p.m. to 8
a.m. in STZ rats was documented by a decrease in glycemia at time zero of the IPGTT, a decrease in the total glycemic AUC during the IPGTT, a lower plasma D-glucose concentration at sacrifice after overnight starvation, and a trend towards a higher plasma insulin concentration and insulinogenic index and a lower HOMA index at sacrifice. For the latter three variables, the geometric means of the relevant variable (plasma insulin concentration, insulinogenic index, and inverse of HOMA index) yielded a significant difference (P
< 0.025) between NF (100.0 ± 6.9%; n
= 15) and IF (164.4 ± 29.5%; n
= 18) STZ rats. Furthermore, no significant adverse effect of intermittent fasting (P
> 0.49) was observed in terms of the changes in body weight of the STZ rats over the 30 days final experimental period, when comparing NF animals (−12.4 ± 2.8
= 5) to IF rats (−18.8 ± 7.9
A different situation prevailed in the calorie-restricted STZ rats. No statistically significant beneficial effects of caloric restriction in the STZ rats was observed when comparing NF to CR diabetic animals. Moreover, the decrease in body weight observed in the STZ rats during the final 30 days experimental period was 2.5 to 3.8 times higher (P < 0.001) in CR rats than in IF and NF animals, respectively.
Even in control rats, the gain in body weight was much lower in CR animals than in IF ones (). This coincided with lower mean values for the plasma insulin concentration, insulinogenic index, and HOMA index in IF control rats than in CR control rats examined at sacrifice after overnight starvation (). Thus, for these three variables, the values recorded in IF control rats averaged 71.6 ± 8.6% (n = 15; P < 0.06) of the mean corresponding values found in CR control rats (100.0 ± 11.5%; n = 15). Since such distinctions between IF and CR control rats could not be ascribed to any difference in either food intake or the responsiveness to D-glucose of isolated pancreatic islets incubated in vitro, they suggest a more stressful situation in CR control rats than in IF control animals. To a large extent, a comparable situation may prevail in CR as distinct from IF diabetic animals.
In conclusion, therefore, the present study allows to extend to streptozotocin-induced diabetic rats, the proposal that intermittent fasting exerts a beneficial effect on glucose tolerance [11
]. In our opinion, such a dietary approach merits to be also considered as a possible approach to prevent or minimize, if not correct, disturbances of glucose homeostasis in human subjects.