EXTRACTS OF MORICANDIA FOR THE USE THEREOF IN THE PREVENTION AND TREATMENT OF METABOLIC DISEASES

Abstract

A composition including an extract of Moricandia for the use thereof in the prevention and/or treatment of metabolic diseases related to impaired glucose tolerance and/or insulin resistance. Also a composition including an extract of Moricandia for the use thereof in the prevention of weight gain, in particular of body fat.

Claims

1-15. (canceled)

16. A method of preventing and/or treating a metabolic disease selected from the group consisting of diabetes, metabolic syndrome, diseases related to insulin resistance or deficiency, glucose intolerance, hyperglycemia, obesity, dyslipidemia, hypercholesterolemia, hypertriglyceridemia and oxidative stress, in a subject in need thereof, comprising administering to said subject a composition comprising an extract of Moricandia.

17. The method according to claim 16, wherein Moricandia is Moricandia arvensis.

18. The method according to claim 16, wherein the extract of Moricandia is a decoction and/or a maceration of Moricandia.

19. The method according to claim 16, wherein the extract of Moricandia is a decoction and/or a maceration of Moricandia arvensis.

20. The method according to claim 16, wherein the extract of Moricandia is a decoction of leaves of Moricandia, optionally lyophilized.

21. The method according to claim 16, wherein the extract of Moricandia is a decoction of leaves of Moricandia arvensis, optionally lyophilized

22. The method according to claim 16, in which the metabolic disease is selected from the group consisting of type 2 diabetes, insulin resistance, glucose intolerance and hyperglycemia.

23. The method according to claim 16, in which the metabolic disease is type 2 diabetes.

24. The method according to claim 16, further comprising an extract of Astragalus.

25. The method according to claim 24, wherein Astragalus is Astragalus armatus.

26. The method according to claim 24, wherein the extract of Astragalus is a decoction and/or a maceration of Astragalus.

27. The method according to claim 24, wherein the extract of Astragalus is a decoction and/or a maceration of Astragalus armatus.

28. The method according to claim 24, wherein the extract of Astragalus is a maceration of Astragalus roots, optionally lyophilized.

29. The method according to claim 24, wherein the extract of Astragalus is a maceration of Astragalus armatus roots, optionally lyophilized.

30. The method according to claim 24, wherein the extract of Astragalus is a methanolic maceration of Astragalus armatus roots, optionally lyophilized

31. A method of: preventing weight gain in a subject; preventing fat mass gain in a subject; controlling weight gain in a subject; controlling fat mass gain in a subject; stimulating weight loss in a subject; or stimulating fat mass loss in a subject; comprising administering to said subject a composition comprising an extract of Moricandia.

32. The method according to claim 31, wherein Moricandia is Moricandia arvensis.

33. The method according to claim 31, wherein the extract of Moricandia is a decoction and/or a maceration of Moricandia.

34. The method according to claim 31, wherein the extract of Moricandia is a decoction of Moricandia leaves, optionally lyophilized.

35. The method according to claim 31, further comprising an extract of Astragalus, optionally lyophilized.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0176] FIG. 1 is a graph showing the evolution of glycemia during the study protocol in the different groups of rats. Glycemia was measured at the start of the protocol and at the end of each of the first three weeks. The legend for the graph reads as follows:

[0177] STD: rats on a standard diet, i.e., reference group;

[0178] HFHS (“High Fat High Sucrose”): rats on a sugar-rich and fat-rich diet, diabetic after 3 weeks;

[0179] HFHS-MA (“High Fat High Sucrose”): rats on a sugar-rich and fat-rich diet, and treated every morning by gavage with 250 mg/kg of an extract of Moricandia arvensis from the start of the HFHS diet;

[0180] HFHS-AA (“High Fat High Sucrose”): rats on a sugar-rich and fat-rich diet, and treated every morning by gavage with 250 mg/kg of an extract of Astragalus armatus from the start of the HFHS diet.

[0181] **p<0.01 and ****p<0.0001: STD vs HFHS

[0182] #### p<0.001: HFHS vs HFHS+MA and HFHS vs HFHS+AA 2-way ANOVA test.

[0183] FIG. 2 is a histogram showing the basal insulinemia after 3 weeks of the study protocol in the different groups of rats as defined in the legend of FIG. 1.

[0184] ***p<0.001

[0185] ### p<0.001

[0186] $ p<0.05

[0187] 2-way ANOVA test.

[0188] FIG. 3 shows the results of the insulin sensitivity test carried out at the end of the 3 weeks of the study protocol in the different groups of rats as defined in the legend of FIG. 1.

[0189] (A) Graph showing the evolution of glycemia during the test. After 45 min following the insulin injection, while glycemia no longer drops in the HFHS group, the HFHS+MA and AA groups show a glycemia which is not significantly different from the controls and very significantly reduced compared to the untreated HFHS group.

[0190] **p<0.01 and ****p<0.0001: HFHS vs. HFHS-MA

[0191] ## p<0.01 and #### p<0.0001: HFHS vs. STD

[0192] ### p<0.001 and #### p<0.0001: HFHS vs. HFHS-AA 2-way ANOVA test.

[0193] (B) Histogram showing the area under the curves of graph (A). AUC: area under the curve.

[0194] **p<0.01: HFHS vs. STD

[0195] # p<0.05: HFHS vs. HFHS+MA and HFHS+AA vs. HFHS 2-way ANOVA test.

[0196] FIG. 4 shows the results of the glucose tolerance test (2 g/kg orally) carried out at 3 weeks of the study protocol in the different groups of rats as defined in the legend of FIG. 1.

[0197] (A) Graph showing the evolution of glycemia during the test. Animals treated with AA and MA have better glucose tolerance compared to non-treated animals.

[0198] **p<0.01 and ***p<0.001 and ****p<0.0001: HFHS vs. STD

[0199] ## p<0.01 and ### p<0.001 and #### p<0.0001: HFHS-MA vs. HFHS

[0200] ## p<0.01 and #### p<0.0001: HFHS vs. HFHS-AA 2-way ANOVA test.

[0201] (B) Histogram showing the area under the curve (glycemia as a function of time in the range 0-60 minutes during the test). Glucose intolerance observed in HFHS rats is not present when the HFHS diet was combined with MA or AA treatment.

[0202] *p<0.05: HFHS vs. STD

[0203] ## p<0.01: HFHS-MA vs. HFHS and HFHS vs. HFHS-AA 2-way ANOVA test.

[0204] (C) Graph showing the evolution of insulinemia during the OGTT test. Insulin secretion in response to glucose is impaired in HFHS rats. Insulin secretion in response to glucose remains impaired in HFHS rats treated with MA. In HFHS rats treated with AA, insulin secretion in response to glucose is equivalent to controls.

[0205] # p<0.05: HFHS vs. STD

[0206] ## p<0.01: HFHS-MA vs. HFHS and HFHS vs HFHS-AA 2-way ANOVA test.

[0207] FIG. 5 shows the results of the body composition measurement after 3 weeks (1 to 3 days following the 3 weeks of the study protocol) in the different groups of rats.

[0208] STD: rats on a standard diet;

[0209] HFHS (“High Fat High Sucrose”): rats on a sugar-rich and fat-rich diet;

[0210] HFHS-MA (“High Fat High Sucrose”): rats on a sugar-rich and fat-rich diet, and treated every morning by gavage with 250 mg/kg of an extract of Moricandia arvensis.

[0211] (A) Histogram illustrating the proportion of fat mass in % of the total weight; % of body water is not shown. The MA treatment helps preventing fat mass gain during the HFHS diet.

[0212] ***p<0.001: HFHS vs. STD

[0213] # p<0.05: HFHS-MA vs HFHS 2-way ANOVA test.

[0214] (B) Histogram illustrating the proportion of lean body mass. The MA treatment helps preventing the lean mass loss during the HFHS diet.

[0215] **p<0.01: HFHS vs. STD

[0216] # p<0.05: HFHS-MA vs HFHS 2-way ANOVA test.

[0217] FIG. 6 shows the evolution of the differences in weight (delta weight) during the study protocol in the different groups of rats as defined in the legend of the FIG. 1.

[0218] (A) Graph illustrating the change in weight during the study protocol. Animals under the HFHS diet show a greater weight gain than STD control rats. Treatment with MA or AA prevents weight gain in HFHS animals.

[0219] *p<0.05 and **p<0.01 and ****p<0.0001: STD vs. HFHS

[0220] ## p<0.01 and #### p<0.001: HFHS vs HFHS+MA

[0221] ## p<0.01 and ### p<0.001: HFHS vs. HFHS+AA 2-way ANOVA test.

[0222] (B) Histogram illustrating weight gain during the study protocol. The HFHS animals treated with MA or AA show an identical weight gain to the STD control rats and significantly lower than the HFHS rats without treatment.

[0223] *p<0.05 and **p<0.01: STD vs. HFHS

[0224] ## p<0.01: HFHS vs. HFHS+MA and HFHS vs. HFHS+AA 2-way ANOVA test.

[0225] FIG. 7 is a histogram showing the insulin secretion capacity of primary islets of Langerhans from C57Bl6 mice cultured for 72 hours under the following conditions:

[0226] control: culture medium alone;

[0227] palmitate: culture medium+0.5 mM palmitate (lipotoxic conditions mimicking the environment of T2D);

[0228] palmitate+FM50 (M. arvensis 50 mg): culture medium+0.5 mM palmitate+50 μg/mL of Moricandia arvensis leaf extract;

[0229] palmitate+FM100 (M. arvensis 100 mg): culture medium+0.5 mM palmitate+100 μg/mL of Moricandia arvensis leaf extract;

[0230] palmitate+RA50 (A. armatus 50 mg): culture medium+0.5 mM palmitate+50 μg/mL of Astragalus armatus root extract;

[0231] palmitate+RA100 (A. armatus 100 mg): culture medium+0.5 mM palmitate+100 μg/mL of Astragalus armatus root extract.

[0232] Insulin secretion was measured in vitro either under basal conditions (non-stimulating glucose concentration: 2.8 mM) or after stimulation (16.7 mM glucose) at the end of the predefined culture conditions. Treatment with an extract of Moricandia arvensis or of Astragalus armatus in “palmitate” condition restores insulin secretion similar to that of islets in normal condition.

[0233] *p<0.05: control vs. palmitate

[0234] ## p<0.01: palmitate vs. P+FM100

[0235] # p<0.05: palmitate vs. P+RA100

[0236] FIG. 8 is a graph showing the evolution of glycemia during the study protocol in the different groups of ZDF rats. Follow-up is done at the end of each week for 12 weeks.

[0237] ZDF Lean (fa/+): control, heterozygous for the recessive mutation or homozygous for the wild-type leptin receptor allele, i.e., reference group.

[0238] ZDF DB (fa/fa): homozygous for the leptin receptor mutation. The animals are obese, insulin-resistant and then irreversibly diabetic.

[0239] ZDF DB (fa/fa)+MA: homozygous for the mutation of the leptin receptor, treated by gavage with 250 mg/kg of a Moricandia arvensis extract prepared as described in FIG. 1 above.

[0240] *p<0.05 and ****p<0.001: ZDF DB vs. ZDF lean

[0241] # p<0.05 and ## p<0.01 and ### p<0.001 and #### p<0.0001: ZDF DB+MA vs. ZDF DB

[0242] Two-way ANOVA test.

[0243] FIG. 9 is a histogram showing the evolution of insulinemia over time (integration of the area under the curve) during the first 8 weeks of the study protocol in the different groups of rats as defined in the legend of the FIG. 8.

[0244] **p<0.01 and ***p<0.001: ZDF DB vs. ZDF lean

[0245] Two-way ANOVA test.

[0246] FIG. 10 shows the results of the insulin sensitivity test (1 U/kg) carried out after 8 weeks of the study protocol in the different groups of rats as defined in the legend of FIG. 8.

[0247] (A) Graph showing the evolution of glycemia during the test. Both groups of ZDF DB and ZDF DB+MA diabetic rats were hyperglycemic compared to the ZDF lean control group during the first 45 minutes following insulin injection. The untreated ZDF DB group is insulin-resistant throughout the test compared to the control ZDF group, whereas in the group treated with MA (ZDF DB+MA), insulin resistance gradually disappears (significant improvement after 60 minutes, to become insignificant compared to the ZDF lean control group). In the interval between 75 and 120 minutes, insulin sensitivity becomes identical between the ZDF DB+MA and ZDF lean groups, thus allowing a complete restoration of glycemia in response to exogenous insulin.

[0248] ****p<0.001: ZDF DB vs. ZDF lean

[0249] **p<0.01 and ****p<0.001: ZDF DB+MA vs. ZDF lean

[0250] Two-way ANOVA test.

[0251] (B) Area under the curve integrating glycemia during the first 60 minutes of the test.

[0252] ZDF DB+MA rats have a lower glycemic excursion than untreated ZDF DB rats but higher than ZDF lean rats.

[0253] **p<0.01: ZDF DB+MA vs. ZDF lean

[0254] ****p<0.0001: ZDF DB vs. ZDF lean

[0255] ## p<0.01: ZDF DB+MA vs. ZDF DB

[0256] Two-way ANOVA test.

[0257] FIG. 11 shows the results of the glucose tolerance test (2 g/kg) carried out after 8 weeks of the study protocol in the different groups of rats as defined in the legend of FIG. 8.

[0258] (A) Graph showing the evolution of glycemia during the test. Glycemia is significantly higher in ZDF DB rats than in ZDF+MA rats.

[0259] **p<0.01 and ****p<0.001: ZDF DB+MA vs. ZDF DB

[0260] ***p<0.001: ZDF DB vs. ZDF lean

[0261] Two-way ANOVA test.

[0262] (B) Histograms illustrating the area under the glycemia trend curves as a function of time in the range 0-60 minutes during the test.

[0263] *p<0.05: ZDF DB+MA vs. ZDF-DB

[0264] ****p<0.001: ZDF DB vs. ZDF lean

[0265] #### p<0.001: ZDF DB+MA vs. ZDF lean

[0266] Two-way ANOVA test.

[0267] FIG. 12 shows in histogram form the lipid balance (total cholesterol and triglycerides) in the 3 groups of rats as defined in the legend of FIG. 8. The levels of total cholesterol (CHOL), triglycerides (TRIG), low density lipoprotein cholesterol (LDLC) and high-density lipoprotein cholesterol (HDLC) are indicated.

[0268] ****p<0.0001: ZDF DB vs. ZDF lean

[0269] # p<0.05 and ### p<0.001: ZDF DB+MA vs. ZDF DB

[0270] Two-way ANOVA test.

EXAMPLES

[0271] The present invention will be better understood in the light of the following examples which illustrate the invention in a non-limitative manner

Example 1

Harvesting and Preparation of Extracts of Moricandia arvensis and Astragalus armatus

[0272] An extract of Moricandia arvensis was prepared by decoction, by immersing 100 g of dried and powdered leaves in 1 liter of boiling water for 15 to 20 minutes. After filtration of the decoction, the extract was lyophilized. The lyophilizate was then stored at 4° C.

[0273] An extract of Astragalus armatus was prepared by maceration, by immersing 100 g of dried and powdered roots in 1 liter of methanol for 15 days with stirring. After filtration of the macerate, methanol was evaporated under reduced pressure with a rotary evaporator at 40° C. until a dry extract (powder) was obtained. The dry extract was then stored at 4° C.

Example 2

Pharmacological Effects of Extracts of Moricandia arvensis and Astragalus armatus in a Development Model of Type 2 Diabetes

[0274] Materials and Methods

[0275] Animals and Study Protocol

[0276] Wistar rats (Charles River laboratories, USA) were divided into 3 groups for a study protocol lasting 26 days: [0277] the rats of group 1 followed a standard diet (SAFE laboratories, France); [0278] the rats of group 2 were fed an HFHS diet (“High Fat High Sucrose” - reference H235, SAFE laboratories, France) which leads to symptoms of type 2 diabetes; [0279] the rats of group 3 were fed an HFHS diet and were treated every morning by gavage with 250 mg/kg with an extract of Moricandia arvensis (HFHS-MA) or of Astragalus armatus (HFHS-AA) prepared as described in Example 1 above.

[0280] Glycemia was measured at the start of the protocol (day 0), at the end of week 1 (day 7), at the end of week 2 (day 14) and at the end of week 3 (day 21). Several tests were then carried out in a period of 5 days at the end of the protocol (days 22 to 26—2-day rest interval between each of the tests). Basal insulinemia was measured. An oral glucose tolerance test, OGTT (oral hyperglycemia −2 g/kg) was performed. Glycemia and insulinemia were measured during this test. An insulin sensitivity test (intraperitoneally) (1 U/kg) was also performed. The body composition was analyzed by magnetic resonance imaging (EchoMRI) for groups 1 and 2, and for animals of group 3 treated with an extract of Moricandia arvensis on day 26 of the study protocol, after the end of the tests. The rats were weighed on days 0, 5, 9, 15, 18, 23, 25 and 26 of the study protocol.

[0281] Statistical Analysis

[0282] The values of the means±standard error of the mean (SEM), during and/or at the end of the treatment, are presented. The size is n=8 rats/group, except for the measurement of basal glycemia and insulinemia for which n=9 to 11. Statistical calculations to assess the differences between the groups (significance level: p<0.05) were performed by a 2-way ANOVA analysis of variance (time/treatment effect). Statistical analyzes were performed using Prism software. Significant differences are shown in the figures.

[0283] Results

[0284] Treatment with an extract of Moricandia arvensis or an extract of Astragaius armatus prevents hyperglycemia observed in untreated animals fed the HFHS diet (FIG. 1 and Table 1).

TABLE-US-00001 TABLE 1 Glycemia at the end of week 3 (day 21) for the different groups. Glycemia Glycemia Group (mM ± SEM) (g/L ± SEM) STD 6.17 ± 0.17 1.10 ± 0.03 HFHS 7.98 ± 0.25 1.43 ± 0.04 HFHS-MA 6.52 ± 0.16 1.17 ± 0.03 HFHS-AA 6.16 ± 0.15 1.10 ± 0.03

[0285] The measurement of basal insulinemia at the end of the protocol (FIG. 2 and Table 2) and the results of the insulin sensitivity test (FIGS. 3A and 3B; Tables 3 and 4) show that the treatment with an extract of Moricandia arvensis or an extract of Astragalus armatus prevents the development of insulin resistance, seen in untreated animals fed the HFHS diet.

TABLE-US-00002 TABLE 2 Basal insulinemia at the end of the protocol for the different groups. Insulin levels Group (μU/mL ± SEM) STD 52.0 ± 0.16 HFHS 62.2 ± 2.00 HFHS-MA 53.4 ± 2.6  HFHS-AA 42.7 ± 4.4 

TABLE-US-00003 TABLE 3 Glycemia at 120 minutes after the insulin injection during the insulin sensitivity test for the different groups. Glycemia Group (μU/mL ± SEM) STD 52.0 ± 0.16 HFHS 62.2 ± 2.00 HFHS-MA 53.4 ± 2.6  HFHS-AA 42.7 ± 4.4 

TABLE-US-00004 TABLE 4 Area under the glycemia trend curves during the insulin sensitivity test. Area Under Curve Group (arbitrary surface unit ± SEM) STD 6951 ± 351 HFHS 9965 ± 654 HFHS-MA 7634 ± 375 HFHS-AA 7479 ± 493

[0286] The glucose tolerance test shows that treatment with an extract of Moricandia arvensis or an extract of Astragalus armatus prevents the development of glucose intolerance, observed in untreated animals fed with the HFHS diet (FIGS. 4A and 4B; Tables 5 and 6). Measurement of insulinemia during the glucose tolerance test shows that the insulin level does not change in animals of group 3 treated with an extract of Moricandia arvensis (FIG. 4C; Table 7). This observation suggests that the effect of Moricandia arvensis extract on glycemia is mediated by the preservation of insulin sensitivity. In animals treated with an extract of Astragalus armatus, insulinemia is equivalent to that measured in the control group, fed a standard diet (FIG. 4C). This finding indicates that treatment with an extract of Astragalus armatus, in addition to its effect on insulin sensitivity, also prevents the decrease in insulin secretion capacity observed in untreated animals fed the HFHS diet.

TABLE-US-00005 TABLE 5 Glycemia during the glucose tolerance test. Glycemia (mg/dl. ± SEM) Time Group 15 minutes 45 minutes 60 minutes 90 minutes 120 minutes STD — 144.4 ± 5.5 141.8 ± 3.9 135.2 ± 1.8 — HFHS 180.3 ± 5.9 173.9 ± 3.5 175.9 ± 5.1 160.0 ± 3.1 143.5 ± 4.9 HFHS-MA 154.4 ± 5.5 150.3 ± 6.0 146.0 ± 4.7 129.3 ± 4.9 118.0 ± 3.1 HFHS-AA — 150.6 ± 4.5 138.3 ± 3.9 128.1 ± 3.8 116.5 ± 1.6

TABLE-US-00006 TABLE 6 Area under the glycemia trend curves during the glucose tolerance test in the 0-60-minute time interval. Area under the curve Group (arbitrary surface unit ± SEM) STD 8882 ± 273 HFHS 10100 ± 92  HFHS-MA 8686 ± 284 HFHS-AA 8899 ± 230

TABLE-US-00007 TABLE 7 Insulinemia during the glucose tolerance test at time +15 minutes. Group Insulin levels (μU/mL ± SEM) STD 67.2 ± 5.1 HFHS 46.3 ± 5.1 HFHS-MA 45.1 ± 5.1 HFHS-AA 74.4 ± 4.5

[0287] Analysis of body composition shows that treatment with an extract of Moricandia arvensis prevents the increase in the proportion of body fat mass (and the decrease in the proportion of associated lean body mass) observed in untreated animals fed the HFHS diet (FIGS. 5A and 5B; Table 8). This finding indicates a potential mechanism related to the development of body fat mass underlying the preservation of insulin sensitivity in animals treated with an extract of Moricandia arvensis.

TABLE-US-00008 TABLE 8 Analysis of body composition. Fat mass Lean body mass (% of total weight (% of total weight Group without water ± SEM) without water ± SEM) STD 13.7 ± 0.7 72.5 ± 0.7 HFHS 20.0 ± 1.1 66.9 ± 1.0 HFHS-MA 16.3 ± 0.9 71.7 ± 1.9

[0288] The weighing of the animals in the 3 groups during the study protocol shows that the treatment with an extract of Moricandia arvensis or an extract of Astragalus armatus prevents weight gain observed in untreated animals fed with the HFHS diet (FIG. 6 and Table 9).

TABLE-US-00009 TABLE 9 Evolution of the difference in weight on days 9, 18, 23 and 26. Weight delta (g ± SEM) Day Group 9 18 23 STD 26.8 ± 1.7 40.4 ± 2.2 65.1 ± 3.0 HFHS 45.5 ± 5.0 81.0 ± 5.7 101.6 ± 7.8  HFHS-MA 29.6 ± 3.3 52.1 ± 4.4 68.4 ± 7.0 HFHS-AA 28.4 ± 1.8 52.9 ± 2.7 79.0 ± 2.9

[0289] Conclusion

[0290] These results, taken as a whole, show that treatment with an extract of Moricandia arvensis or an extract of Astragalus armatus prevents hyperglycemia, insulin resistance, glucose intolerance and the weight gain observed in a model developing symptoms of type 2 diabetes.

[0291] In the case of treatment with an extract of Astragalus armatus, it appears that this also preserves the insulin secretion capacity, which is reduced when the animals are fed the HFHS diet.

Example 3

Influence of Extracts of Moricandia arvensis and Astragalus armatus on the Capacity for Insulin Secretion by Pancreatic Islets of Rats in Primary Culture

[0292] Materials and Methods

[0293] Pancreatic islets from C57Bl6 mice were isolated by size, distributed into culture wells (taking care to respect an equivalent distribution in each well in terms of size) and cultured under normal condition (control) or under lipotoxic condition in the presence of palmitate: [0294] [Control]: culture medium alone; [0295] [Palmitate]: culture medium+0.5 mM palmitate (saturated fatty acid mimicking diabetic conditions—DT2); [0296] [Palmitate+FM50]: culture medium+0.5 mM palmitate+50 μg/mL of extract of Moricandia arvensis; [0297] [Palmitate+FM100]: culture medium+0.5 mM palmitate+100 μg/mL of extract of Moricandia arvensis; [0298] [Palmitate+RA50]: culture medium+0.5 mM palmitate+50 μg/mL of extract of Astragalus armatus; [0299] [Palmitate+RA100]: culture medium+0.5 mM palmitate+100 μg/mL of extract of Astragalus armatus;

[0300] Extracts of Moricandia arvensis and Astragalus armatus were prepared as described in Example 1 above.

[0301] After 72 hours of culture, the insulin secretion capacity of the pancreatic islets was tested by measuring the concentration of insulin before and after stimulation by an increase in glucose concentration in the medium from 2.8 mM (basal condition) to 16.7 mM (stimulated condition).

[0302] Results

[0303] The basal secretion capacity and after stimulation of the pancreatic islets, reduced in the presence of palmitate, is largely preserved by the presence of an extract of Moricandia arvensis or Astragalus armatus in the medium (FIG. 7 and Table 10).

TABLE-US-00010 TABLE 10 Insulin secretion by the pancreatic islets. Insulin secretion (ng/islet/h ± SEM) Simulation Culture condition No Yes Control 0.126 ± 0.010 2.869 ± 0.066 Palmitate 0.084 ± 0.016  1.845 ± 0.0183 Palmitate + FM50 0.118 ± 0.014 2.525 ± 0.280 Palmitate + FM100 0.141 ± 0.002 2.430 ± 0.255 Palmitate + RA50 0.117 ± 0.016 2.279 ± 0.163 Palmitate + RA100 0.126 ± 0.011 2.349 ± 0.324

[0304] Conclusion

[0305] These results indicate a protective effect of extracts of Moricandia arvensis and Astragalus armatus on the capacity of insulin secretion by pancreatic cells, and thus the onset of type 2 diabetes in the event of a sugar-rich and fat-rich diet.

Example 4

Curative Effects of MA in a Genetic Model of Type 2 Diabetes (Zücker ZDF Rat)

[0306] Materials and Methods

[0307] Animals and Study Protocol

[0308] The anti-diabetic effects of MA extracts were tested in a severe T2D model: the ZDF rat (Zücker diabetic fatty rat), a genetic model which exhibits a recessive mutation of the leptin receptor (only animals carrying the mutation on both alleles develop obesity and T2D). This total resistance to leptin leads to overeating, massive obesity and insulin resistance which are irreversible: diabetes sets in and continues to increase with age. First, hyperinsulinemia develops at weaning (at 3 weeks of age) to compensate for tissue resistance, but the endocrine pancreas becomes depleted and the animals quickly become diabetic, as their insulin secretion declines and eventually breaks down.

[0309] It is in this model that the extracts of MA were tested, under the same conditions as for the preventive effects of the preceding Examples (250 mg/kg, by daily gavage, for 8 to 13 weeks).

[0310] The 6-week-old animals were divided into 3 groups for a 13-week study protocol: [0311] ZDF Lean (fa/+): control, heterozygous for the recessive mutation or homozygous for the wild-type leptin receptor allele. This is the normal reference group. [0312] ZDF DB (fa/fa): homozygous for the leptin receptor mutation. The animals are obese, insulin resistant and then irreversibly diabetic. [0313] ZDF DB (fa/fa)+MA: homozygous for the leptin receptor mutation, treated by gavage with 250 mg/kg of an extract of Moricandia arvensis prepared as described in Example 1 above.

[0314] The animals are on a normal diet (SAFE). The various measurements and tests were carried out during the 8 to 13 weeks.

[0315] Statistical Analysis

[0316] The size per group is n=6. Results are presented as the mean±standard error of the mean (SEM). Statistical analyzes are analyzes of variance (ANOVA), with a significance level at p<0.05. Significant differences are shown in the figures.

[0317] Results

[0318] Treatment with an extract of Moricandia arvensis decreases hyperglycemia observed from week 3 in untreated leptin receptor mutation homozygotes (FIG. 8 and Table 11).

TABLE-US-00011 TABLE 11 Glycemia at week 11 of the study protocol. Group Glycemia (mg/dL ± SEM) ZDF lean 116.5 ± 3.6  ZDF DB 525.8 ± 20.2 ZDF DB + MA 454.1 ± 30.3

[0319] The measurement of basal insulinemia during the first 8 weeks of the protocol (FIG. 9 and Table 12) and the results of the insulin sensitivity test (FIG. 10A and 10B; Table 13) show that treatment with an extract of Moricandia arvensis treats the insulin resistance observed in untreated leptin receptor mutant homozygotes.

TABLE-US-00012 TABLE 12 Area under the basal insulinemia trend curves during the first 8 weeks of the protocol. Area under the curve Group (arbitrary surface unit ± SEM) ZDF lean  8.11 ± 0.17 ZDF DB 13.76 ± 0.76 ZDF DB + MA 12.03 ± 0.92

TABLE-US-00013 TABLE 13 Area under the glycemia trend curves during the first 60 minutes of the insulin sensitivity test. Area under the curve Group (arbitrary surface unit ± SEM) ZDF lean 243.5 ± 33.9 ZDF DB 1424.0 ± 119.9 ZDF DB + MA 964.9 ± 71.4

[0320] The glucose tolerance test shows that treatment with an extract of Moricandia arvensis makes it possible to reduce the glucose intolerance observed in untreated leptin receptor mutant homozygotes (FIGS. 11A and 11B; Table 14).

TABLE-US-00014 TABLE 14 Area under the glycemia trend curves in the first 60 minutes of the glucose tolerance test. Area under the curve Group (arbitrary surface unit ± SEM) ZDF lean 1183.8 ± 8.5  ZDF DB 4371.0 ± 93.2 ZDF DB + MA 3472.8 ± 67.6

[0321] The lipid balance shows that the treatment with an extract of Moricandia arvensis makes it possible to reduce the level of cholesterol, triglycerides and high-density lipoprotein cholesterol (HDLC) abnormally high in untreated leptin receptor mutant homozygotes (FIG. 12 and Table 15). Given the inflammatory role of lipids and the role of inflammation in the development of insulin resistance, one of the potential effects of Moricandia arvensis could be to improve the lipid balance.

TABLE-US-00015 TABLE 15 Lipid balance. Total cholesterol Triglycerides Group (mmol/L ± SEM) (mmol/L ± SEM) ZDF lean 2.0 ± 0.1 1.4 ± 0.1 ZDF DB 4.4 ± 0.1 5.1 ± 0.4 ZDF DB + MA 3.9 ± 0.1 4.0 ± 0.3

[0322] Conclusions

[0323] These results, taken as a whole, show that the administration of an extract of Moricandia arvensis can curatively treat type 2 diabetes in this preclinical model.

Example 5

Pharmacological Effects of a Combination of Extracts of Moricandia arvensis and Astragalus armatus in a Developmental Model of Type 2 Diabetes

[0324] Materials and Methods

[0325] Animals and Study Protocol

[0326] Wistar rats (Charles River laboratories, USA) are divided into 3 groups for a study protocol lasting 26 days: [0327] the rats of group 1 follow a standard diet (SAFE laboratories, France); [0328] the rats of group 2 are fed an HFHS diet (“High Fat High Sucrose”—reference H235, SAFE laboratories, France) which leads to the symptoms of type 2 diabetes; [0329] the rats of group 3 were fed an HFHS diet, and were treated every morning by gavage with 250 mg/kg of a combination of extracts of Moricandia arvensis and Astragalus armatus (HFHS−MA+AA).

[0330] The glycemia is measured at the start of the protocol (day 0), at the end of week 1 (day 7), at the end of week 2 (day 14) and at the end of week 3 (day 21). Several tests are then carried out in a period of 5 days at the end of the protocol (days 22 to 26 −2-day rest interval between each of the tests). Basal insulinemia is measured. An oral glucose tolerance test, OGTT (oral hyperglycemia −2 g/kg) is performed. Glycemia and insulinemia are measured during this test. An insulin sensitivity test (intraperitoneally) (1 U/kg) is also performed. Body composition is analyzed by magnetic resonance imaging (EchoMRI) for groups 1 and 2, and for group 3 animals treated with a combination of extracts of Moricandia arvensis and Astragalus armatus on day 26 of the study protocol, after the end of the tests. Rats are weighed on days 0, 5, 9, 15, 18, 23, 25 and 26 of the study protocol.

Example 6

Influence of a Combination of Extracts of Moricandia arvensis and Astragalus armatus on the Capacity for Insulin Secretion by Pancreatic Islets of Rats in Primary Culture

[0331] Materials and Methods

[0332] Pancreatic islets from C57Bl6 mice were isolated by size, distributed into culture wells (taking care to respect an equivalent distribution in each well in terms of size) and cultured under normal condition (control) or under lipotoxic condition in the presence of palmitate: [0333] [Control]: culture medium alone; [0334] [Palmitate]: culture medium+0.5 mM palmitate (saturated fatty acid mimicking diabetic conditions - DT2); [0335] [Palmitate+FM50]: culture medium+0.5 mM palmitate+50 μg/mL of extract of Moricandia arvensis; [0336] [Palmitate+FM100]: culture medium+0.5 mM palmitate+100 μg/mL of extract of Moricandia arvensis; [0337] [Palmitate+RA50]: culture medium+0.5 mM palmitate+50 μg/mL of extract of Astragalus armatus; [0338] [Palmitate+RA100]: culture medium+0.5 mM palmitate+100 μg/mL of extract of Astragalus armatus;

[0339] Extracts of Moricandia arvensis and Astragalus armatus are prepared as described in Example 1 above.

[0340] After 72 hours of culture, the insulin secretion capacity of the pancreatic islets is tested by measuring the concentration of insulin before and after stimulation by an increase in the concentration of glucose in the medium from 2.8 mM (basal condition) to 16.7 mM (stimulated condition).

Example 7

Curative Effects of a Combination of Extracts of Moricandia arvensis and Astragalus armatus in a Genetic Model of Type 2 Diabetes (Zücker ZDF Rat)

[0341] Materials and Methods

[0342] Animals and Study Protocol

[0343] The anti-diabetic effects of MA extracts are tested in a severe T2D model: the ZDF rat (Zücker diabetic fatty rat), a genetic model which exhibits a recessive mutation of the leptin receptor (only animals carrying the mutation on both alleles develop obesity and T2D). This total resistance to leptin leads to overeating, massive obesity and insulin resistance which are irreversible: diabetes sets in and continues to increase with age. First, hyperinsulinemia develops at weaning (at 3 weeks of age) to compensate for tissue resistance, but the endocrine pancreas becomes depleted and the animals quickly become diabetic, as their insulin secretion declines and eventually breaks down.

[0344] It is in this model that the extracts of Moricandia arvensis and Astragalus armatus are tested, under the same conditions as for the preventive effects of the preceding Examples (250 mg/kg, by daily gavage, for 8 to 13 weeks).

[0345] The 6-week-old animals are divided into 3 groups for a 13-week study protocol: [0346] ZDF Lean (fa/+): control, heterozygous for the recessive mutation or homozygous for the wild-type leptin receptor allele. This is the normal reference group. [0347] ZDF DB (fa/fa): homozygous for the leptin receptor mutation. The animals are obese, insulin resistant and then irreversibly diabetic. [0348] ZDF DB (fa/fa)+MA: homozygous for the leptin receptor mutation, treated by gavage with 250 mg/kg of an extract of Moricandia arvensis prepared as described in Example 1 above.

[0349] The animals are on a normal diet (SAFE). The various measurements and tests are carried out during the 8 to 13 weeks.