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Effect of exercise in combination with dietary nopal and zucchini on chronic and acute glucohomeostasis in genetically obese mice

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Cite this article: Effect of exercise in combination with dietary nopal and zucchini on chronic and acute glucohomeostasis in genetically obese mice. Inte J Expe Bio. 2018; 1(1): 001-005. ABSTRACT Objective: Type 2 Diabetes Mellitus (DM) is a major health problem for which folk medicinals such as nopal, or cactus stems from various species of Opuntia (Cactaceae), are often used as an inexpensive palliative dietary supplement. We sought to determine whether exercise and nopal, independently or interactively, could suppress the normal increase in blood glucose in genetically obese mice. Methods: We tested the effects of nopal (experimental) and zucchini (control) supplemented diets, with and without exercise, on chronic and acute blood glucose levels in genetically obese (C57BL/6JLep ob/ob) mice, which mimic type 2 DM. Results: A normal developmental increase in blood glucose was mitigated by nopal and exercise. A control vegetable, zucchini, was less effective in controlling blood glucose. Neither exercise nor vegetable supplementation suppressed the hyperglycemic response to an acute glucose challenge. Conclusion: These results provide evidence that a nopal-supplemented diet combined with aerobic exercise mitigates chronic but not acute hyperglycemia in a genetically obese mouse model of type 2 DM.
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Effect of exercise in combination with dietary nopal and zucchini on chronic and acute glucohomeostasis in genetically
obese mice
Zoë F Christian¹
,2
, Patrick D Craig¹
,2
, Keith Rayburn
3
and Louis N Irwin
2*
¹Program in Physical Therapy
²Department of Biological Sciences, University of Texas at El Paso, TX 79968, USA
3Family Practice Clinic, Gilroy, CA 95020
*Corresponding Author:
Louis Irwin, 1551 Larimer Street, #2703, Denver, CO 80202, USA; Fax: 303-623-4317; E-mail: lirwin@utep.edu
Cite this article:
Effect of exercise in combination with dietary nopal and zucchini on chronic and acute glucohomeostasis in genetically obese mice.
Inte J Expe Bio. 2018; 1(1): 001-005.
Submitted: 29 August 2018; Approved: 04 September 2018; Published: 05 September 2018
ABSTRACT
Objective: Type 2 Diabetes Mellitus (DM) is a major health problem for which folk medicinals such as nopal, or cactus stems from
various species of Opuntia (Cactaceae), are often used as an inexpensive palliative dietary supplement. We sought to determine whether
exercise and nopal, independently or interactively, could suppress the normal increase in blood glucose in genetically obese mice.
Methods: We tested the effects of nopal (experimental) and zucchini (control) supplemented diets, with and without exercise, on
chronic and acute blood glucose levels in genetically obese (C57BL/6JLepob/ob ) mice, which mimic type 2 DM.
Results: A normal developmental increase in blood glucose was mitigated by nopal and exercise. A control vegetable, zucchini, was
less effective in controlling blood glucose. Neither exercise nor vegetable supplementation suppressed the hyperglycemic response to
an acute glucose challenge.
Conclusion: These results provide evidence that a nopal-supplemented diet combined with aerobic exercise mitigates chronic but
not acute hyperglycemia in a genetically obese mouse model of type 2 DM.
Keywords: Hyperglycemia; Exercise; Diabetes Mellitus Type 2; Nopal
INTRODUCTION
Type 2 (adult onset, or non-insulin dependent) Diabetes
Mellitus [DM], is a disorder of carbohydrate metabolism involv-
ing hyperglycemia that can lead to disabling or life-threatening
conditions such as kidney dysfunction [1], peripheral neuropathy
[2], retinopathy [3], and coronary artery disease [4]. Type 2 DM,
is a particularly common correlate of obesity, and dispropor-

Asian ancestry [5]. It tends to be linked especially to poorer diets
and the inability to obtain affordable medication. The traditional
treatment for hyperglycemia is an appropriate combination of
diet, exercise, and medication [6,7]. Folk medicinals are often
used to supplement or replace these treatments in some patient
populations [8,9].
Stems of the prickly pear cactus commonly known as nopal,
including various species of Opuntia (Cactaceae), are thought to

effects [10]. In Mexico and the Southwestern United States, nopal
is frequently taken as a dietary supplement in the belief that
it helps control hyperglycemia [9,11-13], both chronically and
acutely. While some studies indicate that nopal may have hypogly-
cemic properties [10,14-17], these effects have not been observed
in others [18,19]. Animal studies have likewise been inconclu-
sive, with nopal appearing to be more effective in diabetic than in
normoglycemic animals [20-26]. Exercise has long been part of the
treatment regimen for obese and diabetic patients, in view of its

insulin binding and sensitivity [27], lower insulin requirements
[28], improved blood glucose control [29], improved maximum ox-
ygen uptake [30], and a decrease in stress that promotes a sense of
general well-being [31]. Since exercise is an adjunct with dietary
-
mentation coupled with an aerobic exercise regimen warrants
investigation. The combination of folk remedies such as nopal
with exercise could play a major role in the management of obesity

In preliminary experiments we found dietary restriction to be
effective in controlling the dramatic hyperglycemia of genetically
obese C57Bl/6JLepob/ob mice (hereafter, ob/ob mice), suggesting
that this animal could provide a reliable model for investigating
the metabolic basis of obesity and type 2 DM. In this report,
-
dependently and in combination, mitigate the rise in chronic blood
glucose levels characteristic of the ob/ob mouse. We also present
evidence, however, that the same treatment does not attenuate the
rise in blood glucose in response to an acute glucose challenge.
Materials and Methods
Experimental Design and Animals
This study was designed to test the hypothesis that exercise
and vegetable dietary supplements suppress both chronic and
acute hyperglycemia in genetically obese mice. Forty-eight male
C57Bl/6JLepob/ob mice were obtained from the Jackson Labs (Bar
Harbor, ME) at 5 weeks of age. They were housed individually
in polycarbonate cages with about one inch of soft bedding in a
temperature-controlled animal room (~24C) under a constant
(12L:12D) photoperiod.
Half the mice were exercised (E) by forced swimming trials for
20 min three times per week, while the other half were sedentary
(S) in their home cages throughout the experiment. The diets
consisted of lab chow alone (C) or lab chow in combination with
either fresh zucchini (Z) or nopal (P) slices. A 2 X 3 factorial
design therefore generated six groups (CE, ZE, PE, CS, ZS, PS) con-
sisting of 8 mice each. Body weight and blood glucose levels were
measured as dependent variables.
Animal procurement, housing, and experimental treatment
were approved by the University of Texas at El Paso Institutional
Animal Care and Use Committee, in accordance with the NIH Guide
to the Use and Care of Animals.
Diet and Feeding Schedule
Animals were fed PurinaLab Chow and watered ad lib. Chow
remaining was weighed at weekly intervals to the nearest 0.1 g for
calculation of daily consumption. Fresh nopal, Opuntia sp., and
zucchini, Cucurbita pepo (Cucurbitaceae), were obtained from a
local market. Vegetable slices weighing ~ 2-3 g each were soaked
in a fresh 0.1% saccharine solution for ~ 10 min to provide a
non-caloric inducement for the mice to eat them and placed in the
cages on top of the bedding daily. The following day, the remain-
ing vegetables were weighed to the nearest 0.1 g, and corrected
for dehydration based on loss of wet weight by control nopal and
zucchini slices.
Exercise
Mice were exercised by being placed in a tank of warm
International Journal of Experimental Biology Research Article
(32-35o C) water for a period of sustained swimming, three morn-
ings (0900 – 1100) per week. Because of their obesity, mice were

moderately increase the activity level of these severely hypoactive
animals without exhausting them or stressing them more than
necessary, in accordance with principles of humane animal use,
and to reasonably equate with levels of exercise in humans. Blood
samples were not collected on exercise days. Sedentary mice were
left in their home cages throughout the experiment.
Chronic Blood Glucose
Blood glucose levels were determined at the beginning (week
0) and end (week 3) of the experiment. Mice were fasted for 4h
from 1000 to 1400. Blood samples were then collected from a
cut at the distal end of the tail. One or two drops of blood were
immediately assessed with ONE TOUCHglucose test strips in a re-

meter in mg/dl.
Acute Glucose Tolerance
Mice were tested for acute glucose tolerance at the beginning
and end of the experiment. Between 1400 and 1430, mice received
an intraperitoneal injection of a 32 mg/ml solution of glucose
in a volume (ml) equal to 1.4% of their body weight in g. Basal
blood samples were taken prior to glucose loading (0 min), then
subsequent samples were collected at 15, 30, 60, and 90 minutes
post-injection, from a cut at the distal end of the tail. Subsequent
blood samples were obtained by sloughing off existing scab, or
making a fresh cut if necessary. Blood glucose was then assessed
as described above.
Statistical Analysis
Food consumption and the effects of activity level and diet
on chronic blood glucose were tested by two-way ANOVA, and on

Tukey's correction for repeated measures to make individual pair-
wise comparisons.
Results
Overall Food Consumption
Exercised mice consumed 12% less total food than sedentary


df = 2/42, p =0.06) among the different diets. Zucchini added 10%
and nopal added 7% to the total mass of food consumed. Since the
vegetables are very low in caloric content [20,32], the diets were
essentially isocaloric within each activity group. Body weights of

treatment groups over a narrow range (45-46 g).
Effect of Exercise and Diet on Chronic Blood Glucose
Juvenile ob/ob mice show a normal developmental increase in
chronic blood glucose levels with age. Figure 1 shows the change
in chronic blood glucose levels over the three week duration of
the experiment, by treatment group relative to the baseline levels
of blood glucose for all mice at the start of the experiment. The
normal increase in blood glucose was mitigated by both exercise
(F = 11.01, df = 2/41, p = 0.002) and vegetable supplementation

(F = 0.72, df = 2/41, p = 0.49) indicates that diet and exercise
acted independently and additively. Of the two vegetables, nopal
suppressed hyperglycemia more effectively than zucchini. The
normal rise in blood glucose was reduced 42% (p<0.02) by nopal
in sedentary mice and by 89% (p<0.006) in exercised mice, while
reductions with zucchini were only 13% (p >0.56) in sedentary
mice and 38% (p >0.13) in exercised mice.
Effect of Exercise and Diet on Response to An Acute
Glucose Load
Neither exercise alone nor exercise combined with
either zucchini or nopal were effective in reducing the rise
in blood glucose following the glucose challenge. All groups
showed a similar time course in absolute blood glucose lev-
els over the 90 min period of the glucose challenge, though
sedentary mice fed chow only (Group CS) were consistently higher
than the others (Figure. 2A). The starting blood glucose baseline
was also highest for this group, however. To normalize for the
effect of different baseline glucose levels, the data were plotted
after subtracting the zero time value (Fig. 2B). That plot reveals
that no combination of diet and exercise was effective in mitigating
the rise in blood glucose below that seen in the response of Group
CS to a glucose challenge. Our data, therefore, provide no evidence
for suppression of acute glucose tolerance by exercise or vegetable
supplementation in these mice.
Discussion
Chronic Glucohomeostasis
Our hypothesis that chronic hyperglycemia in genetically obese
mice would be mitigated by exercise and at least one of the vegeta-
ble supplements, nopal, was substantiated.
The effectiveness of exercise in mitigating the progressive
hyperglycemia seen in ob/ob mice is consistent with clinical

obesity and type 2 DM [6]. We have shown that caloric restriction
lowers chronic blood glucose levels in older ob/ob mice, but has
little effect on normal juvenile mice at the same age as the animals
in this study [unpublished data]. Since the exercised mice in this

the exercise effect may be due to lower caloric intake. Whether ex-
ercise acts through a direct metabolic effect or indirectly through
appetite suppression or other mechanisms requires further study,
but our results demonstrate that the ob/ob mouse should be a
useful animal model for analyzing the mechanisms by which exer-
cise helps control hyperglycemia.
The effectiveness of nopal in controlling blood glucose levels
in genetically obese mice is likewise consistent with evidence for

[17]. The suppression of blood glucose levels by nopal contrasted
with the ineffectiveness of zucchini. This observation, plus the fact
that all diets were essentially isocaloric, supports the possibility
that nopal contains a pharmacologically active substance that acts

ob/ob mouse provides an animal model in which these mecha-
nisms can be investigated as well.
Acute Glucohomeostasis
Our hypothesis that acute hyperglycemia following a
glucose challenge in genetically obese mice would also be
mitigated by exercise and vegetable supplements was not
supported. Our failure to detect a suppression of glucose
tolerance with a dietary supplement of nopal contrasts
with reports of some animal studies [22-24], but not others
[20,21]. Whether these differences are attributable to

characteristics of the ob/ob mouse must await further stud-
ies. The lack of clinical evidence for an acute hypoglycemic
effect by nopal has been noted also in both the fasting state
[19] and in response to a glucose challenge in fasting type
2 DM patients [15]. Until the relevant variables are better

premature to generalize about the effect of nopal on acute
glucose loads.
Clinical Relevance
We have shown that dietary supplements of nopal mitigate
the progressive rise in blood glucose seen in one strain of ge-
netically obese mice, particularly when combined with regular
exercise. While the pathophysiology may well be different between
ob/ob mice and human patients with Type 2 DM, these results
substantiate the importance of including exercise as a component
of the treatment regimen for obesity and type 2 DM, and provide
cy of nopal in controlling chronic
hyperglycemia,

nopal ingestion was restricted to chronic rather than acute effects
in the ob/ob mice. Clinical studies with human patients are need-
ed to determine whether the lack of nopal to mitigate an acute
glucose challenge, as seen in ob/ob mice, is clinically relevant to
humans.
ACKNOWLEDGEMENTS
This work was supported by grants from the Research Centers
at Minority Institutions (#G12-RRO8124), National Center for Re-
search Resources, NIH, and the Howard Hughes Medical Institute.
Z.F.C. received support from a Houston Endowment Fellowship,
and L.N.I. was supported by an endowment from J. Edward and
Helen M. C. Stern. We thank Don C. Miller for technical assistance,
and Dr. Jerry Hunter (deceased) for valuable suggestions and
advice.
Table 1: Food consumption in sedentary and exercised obese mice
with and without vegetable supplementation
Group Activity Food Consumed (g/day)
Chow Zucchini Nopal
CS sedentary 6.31± 0.21 - -
ZS sedentary 7.11 ± 0.27 0.70 ± 0.08 -
PS sedentary 6.73 ± 0.33 -0.41 ± 0.03
CE exercised 5.75 ± 0.18 - -
ZE exercised 6.33 ± 0.21 0.6 ± 0.09 -
PE exercised 5.29 ± 0.44 - 0.43 ± 0.05
Mice were left in their home cages (sedentary) or given 20 min of
aerobic exercise 3 times per week (exercised). Each activity group
was fed lab chow alone, or chow supplemented with zucchini or
nopal strips ad lib. Numbers represent meanss.e.m. (n = 8) for
dry lab chow and for vegetables after correction for loss of water

Mice in all groups varied in body weight over a very narrow range
of 29-30 g at the start and 45-46 g at the end of the experiment.
Figure 1 : Effect of activity and diet on chronic blood glucose
levels in ob/ob mice. Histograms represent means ± s.e.m. for n =
7-8. Mice were left in their home cages without exercise (Sed-
entary) or given 20 min of aerobic swimming 3 times per week
(Exercised). Each activity group was fed lab chow alone, or chow
supplemented with zucchini or nopal strips ad lib. The horizontal

for baseline blood glucose for all mice at the beginning of the 3
week experiment. Note the rise in chronic blood glucose in all mice
except those that were exercised and fed nopal.
Figure 2: Effect of activity and diet on the time course of change in
blood glucose following a glucose challenge . Mice were fed chow
alone (C) or chow supplemented with Zucchini (Z) or nopal (P) for
three weeks, and left sedentary (S) or exercised (E) 3 times/week
as described under “Methods”. After 3 weeks, a baseline level of
blood glucose was measured (0 min), then all mice were injected
intraperitoneally with a glucose solution (32 mg/ml in a volume
[ml] equal to 1.4% of their body weight in g), and blood was sam-
pled at 15, 30, 60, and 90 min post injection for measurement of
blood glucose. Values indicated are means per treatment group (n

curve. (a) Absolute blood glucose levels. Group CS differed signif-
icantly (p < .05) from Groups CE, ZE, and PE at 15 min, and from
Groups ZS and ZE at 30 min. No other pairwise comparisons were

the baseline (0 min) value for each group.
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