Effects of THC-rich Cannabis sativa extract on Biochemical Parameters in Obesity

doi:10.21203/rs.3.rs-5282770/v1

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Effects of THC-rich Cannabis sativa extract on Biochemical Parameters in Obesity

https://doi.org/10.21203/rs.3.rs-5282770/v1

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The excessive fat accumulation is the cause of obesity that leads to systemic inflammation, compromising the functioning of the brain. Therefore, it is necessary to investigate new effective therapeutic approaches to control fat accumulation. Due to Cannabis sativa anti-inflammatory properties, the use of this plant may be a promising strategy. The objective of this study was to evaluate the effects of THC-rich Cannabis sativa extract (CSE) on the biochemical parameters of obese mice brain. Forty-eight male Swiss mice were used; they were fed a normal-fat or high-fat diet for 10 weeks. On the eighth week of the study, those mice were paired into 4 groups: control+vehicle, control+CSE, obesity+vehicle, obesity+CSE; they received 1mL/kg/day of CSE or olive oil until the end of the study. Body weight was assessed weekly. At the end of the experiment, the mesenteric fat was removed and weighed, and the brain structures were collected for biochemical analyses. The outcome of our study demonstrated that obesity led to mitochondrial and deoxyribonucleic acid (DNA) damage, and that treatment with CSE demonstrated to be effective in reversing this damage. This outcome showed an increase in complex I activity in the hypothalamus and complex II in the prefrontal cortex, but the CSE reversed the damage caused by obesity. Furthermore, a reversion of DNA damage caused by obesity in the mice cortex was observed. It was concluded that despite the need for additional investigations, CSE can be a promising alternative for the treatment of obesity and its consequences.

Cannabis sativa

Obesity

Respiratory chain

THC

Obesity is considered an abnormal accumulation of fat that can be harmful to health according to the World Health Organization (WHO) [1]. This disease has grown in recent years, raising the alarm to health professionals and health organizations. In 2022, WHO estimated that 2.5 billion adults were overweight, and out of these 890 million had some degree of obesity [1].

Unwanted weight gain, which can lead to overweight and obesity, has become a significant factor in the global rise of chronic non-communicable diseases. Obesity is now considered a chronic condition with the potential to impact various aspects of health. Adverse effects include disruptions in female puberty and pubertal disorders, cardiovascular diseases, intestinal health issues, bone and joint disorders, as well as type 2 diabetes mellitus (DM2), among others [2–6].

Classified as a low-grade chronic inflammatory disease, obesity has as its main cause the imbalance between high energy consumption and energy expenditure [7, 8]. Excess energy is stored in the form of triglycerides in adipose tissue, resulting in an accumulation of this fat in adipocytes, causing cell hypertrophy and tissue hyperplasia [9].

The growth in adipocyte mass triggers a local inflammatory process by recruiting immune cells and secreting pro-inflammatory cytokines that are activated mainly by localized hypoxia [10]. This inflammation causes an imbalance in the functioning and signaling of the modulating hormones [11] deregulation in food consumption, control of satiety and energy storage, in addition to being associated with an increase in the production of reactive oxygen species (ROS). ROS are a consequence of electron leakage resulting from an imbalance in the functioning of the mitochondrial respiratory chain, especially in complexes I and II. These complexes receive electrons from the Krebs cycle, and when impaired, they transport fewer electrons than they receive to the complex III, favoring this process [12]. Excess ROS are not sufficiently challenged by the body's natural antioxidant system, causing oxidative stress [13].

Oxidative stress is closely linked to the cells protein and lipid structures; thus, it is common to observe DNA damage in obesity, which can result in problems associated with gene transcription and transduction, as well as impairment of cell replication and the healthy renewal of affected tissues [14]. The relationship between obesity and oxidative stress is complex and includes chronic inflammation, free radical production, mitochondrial dysfunction, antioxidant system imbalance, and other metabolic disorders. These elements create a bidirectional interaction, where obesity leads to increased oxidative stress, which in turn contributes to the development of metabolic disorders and obesity-related diseases [15].

Despite being more explored in peripheral structures, the effects of obesity are also perceived in the central nervous system (CNS) [16], especially in the hypothalamus [12], hippocampus [17], prefrontal and striatum [18, 19]. The hypothalamus is the structure involved in hormonal regulation and regulator of energy metabolism [12], the hippocampus is related to memory, cognition and learning [20]; the prefrontal cortex and striatum are involved in the functioning of the reward system [21]. Therefore, the neuroinflammation caused by obesity is associated with dysfunctions of all such functions.

Therefore, the changes caused by obesity require specific treatments, since the first line treatment (reduction in caloric intake and physical activity) is often patient-refractory. Drug treatments are associated with changing lifestyle habits, having weight loss as their main objective. However, only a reduced number of medications for the treatment of obesity is available and is known to cause significant adverse effects. Thus those drugs are not recommended for use for more than two years [22, 23]. Therefore, it is necessary to offer new pharmacological options that are not only capable of causing weight loss, but that also efficiently reverse the metabolic changes caused by fat accumulation. In this connection, Cannabis sativa could be a treatment option for obesity [24].

Cannabis sativa has several components and among them are cannabinoids, which are substances capable of interacting with the CB1 and CB2 receptors in the human body [25]. The most widely studied cannabinoids, considered the main ones are Tetrahydrocannabivarin (THC), with psychotropic activity and high affinity for CB1, and cannabidiol (CBD) with mainly peripheral action (CB2). The literature strongly supports the anti-inflammatory and antioxidant effects of CBD with its CB2 agonist associated with immune cells modulation. However, other substances present in the plant demonstrate anti-inflammatory potential and should be further explored [24, 26–28].

THC is a substance that has high tropism towards the CNS, being a highly lipophilic crystalline compound, which facilitates its entry into the body and its passage through the blood-brain barrier (BBB), thus causing a faster psychoactive effect [27]. In addition to having free access to the CNS, THC has a high antioxidant potential, due to its ability to bind and neutralize free radicals [29]. In addition to its antioxidant potential, the literature demonstrates that THC has some anti-inflammatory effect. Kaddour and collaborators [30] showed that THC was able to reduce the expression of RNAs associated with inflammation and microglial activation. Therefore, THC may play an important role in the treatment of obesity neuroinflammation, as it has greater lipophilicity that helps it reaching the CNS [27].

However, THC alone does not have many satisfactory effects, so the use of the extract is recommended. This indication is justified by the “Entourage Effect” [31], which is the combined action of endocannabinoids together, since THC associated with CBD can enhance their desired effects and reduce their adverse effects, where a real synergistic interaction and a pharmacological amplification effect may arise [31–33].

Starting from the aforementioned understanding of the impacts of obesity on metabolism, inflammation, and oxidative stress, and the potential effects of Cannabis sativa, the present study aimed to investigate the effect of CSE on the biochemical parameters of obese mice brain.

2.1 Animals

A total of 48 male Swiss miceMus musculus species), 40 days old, weighing around 30 to 40g each, were used. The animals had free access to water and food and were maintained in a 12-hour light-dark cycle at a temperature of 23 ± 1°C. The mice were kept in cages housing 3 to 5 animals, all from the same strain, in order to avoid aggressive behaviors and consequent injuries, which could jeopardise the study.

2.2 Extract

The CSE was donated by a patients’ association (Cannabis Without Borders Association) that handmakes the medicine. CSE was administered after dilution in extra virgin olive oil (vehicle) in a proportion of 1% CSE to 99% olive oil using a 30–35ºC water bath.

The CBD and Δ9-THC content was measured using the High Performance Liquid Chromatography (HPLC) method. The quantities obtained were 3.6 mg/mL CBD and 50.2 mg/mL Δ9-THC.

2.3 Study design

Initially, the animals were weighed and paired into a control group (normal fat diet, n = 24) and an obese group (high fat diet, n = 24), and were maintained in each group for ten weeks under obesity induction according to a feeding protocol by Kalupahana et al.[34] and Cintra et al [35]. To monitor the process, body weight was checked weekly. At the beginning of the eighth week, the animals were divided again into 4 groups (n = 12 per group): Control group + vehicle, Control group + CSE, Obese group + vehicle, Obese group + CSE. The animals received CSE or olive oil at a dosage of 1 mL/kg/day, orally, by gavage. At the end of the experiment, the animals were submitted to euthanasia procedure (painless assisted death). After their death the mesenteric fat was removed and weighed and the brain structures (hypothalamus, hippocampus, prefrontal cortex and striatum) were removed for biochemical analyses and the cerebral cortex was isolated for DNA damage analysis (Fig. 1).

The animal feed was purchased from a company specializing in the development and production of standardized diets for animal experiments (PragSoluções Biociências).

2.4 Body weight and visceral fat

The body weight of the animals was checked weekly by weighing the animals individually and the weight was expressed in grams.

2.5 Tissue removal and storage

After the death of the animals, the abdominal cavity was opened and the adipose tissue from the mesenteric region, located along the intestinal tract, was removed and weighed. The animals' brain was also quickly removed and, subsequently, the brain structures of the hypothalamus, hippocampus, prefrontal cortex and striatum were isolated. Afterwards, the samples were stored at -80°C until the biochemical analysis was performed.

2.6 Mitochondrial respiratory chain enzymes activity analysis

The activity of the respiratory chain complexes was measured in the hypothalamus, hippocampus, prefrontal cortex and striatum. The activity of complex I was evaluated using the method described by Cassina and Radi [36] using the NADH-dependent rate of ferrocyanide reduction. A buffer of 100 mM potassium phosphate, 10 mM ferricyanide, 14 mM NADH, 2 mM rotenone and the sample were added to the reaction medium. After adding all the reagents and the sample, spectrophotometer reading was performed during a period of 3 min, (intervals of 1 min.) at 420 nm.

Complex II activity was measured using the method described by [37] by decreasing the absorbance of 2,6-DCIP. The sample was added to an incubation medium containing 62.5 mM potassium phosphate buffer, 250 mM sodium succinate and 0.5 mM 2,6-DCIP. Incubation was carried out for 20 minutes in a 30°C water bath. After incubation, 100 mM sodium azide, 2 mM rotenone and 0.5 mM 2,6-DCIP were added, and a spectrophotometer reading was carried out every 1 min, for 5 min, at 600 nm.

All the analyses of mitochondrial respiratory chain complexes activity was normalized by protein content, determined as described by Lowry et al. [38]. The results of the mitochondrial respiratory chain complexes activity was expressed in nmol/min x mg protein [37].

2.7 DNA damage analysis

To carry out the comet assay, the cerebral cortex was used. The comet assay was carried out as described in a previous study [39]. Cerebral cortex samples were dissected and immersed in refrigerated phosphate-buffered saline (PBS). Then, the cerebral cortex was individually homogenized with the aid of a syringe, moving it back and forth in order to obtain a cell suspension. Cell suspensions (70 µL aliquots) were embedded in low-melting point agarose (LMA) and then distributed into two well-slides coated with normal melting point agarose (1.5 %, /v) and covered with two coverslips. Once the agarose was solidified, the coverslips were removed and the material was placed in a lysis solution (2.5 M NaCl, 100 mM EDTA, 10 mM Tris, pH 10.0-10.5, with fresh addition of Triton X – 100 and 10% DMSO) for at least 1 hour, protected from light and incubated in new alkaline buffer (pH 12.6) for 30 minutes to unfold the DNA. The cells were subjected to an electrophoresis run, with the same buffer, for 20 minutes at 30 volts and with the use of 300 milliamps electric current and were subsequently neutralized with a neutralizing solution. To assess DNA damage, the slides were stained with Syber Gold for 30 minutes and viewed under a fluorescence microscope at 200x magnification.

A total of 100 cells was evaluated per animal (50 cells in each duplicate slide); the cells were evaluated individually, being categorized into five classes, according to the size of the tail (0 for absence of tail, up to 4 for maximum length tail). This creates a Damage Index (DI) and a Damage Frequency (DF in %) for each animal, with the DI varying from zero (100 x 0 = 0; 100 cells were seen completely without damage) to 400 (100 x 4 = 400; 100 cells with maximum damage) and the DF was calculated based on the number of tail-cells versus the number of cells without a tail. International guidelines and recommendations for the Comet Assay have considered the 100-comet visual to be a well-validated assessment method that has high correlation with computer image analysis [40]. Negative and positive controls were used for each electrophoresis test in order to ensure the reliability of the procedure. All slides were coded for blind analysis.

2.8 Statistical analysis

Data analyses were performed using the InStat Statistical Software (GraphPad, La Jolla, CA, USA). Comparisons between experimental groups were performed using analysis of variance (ANOVA) followed by Tukey's post hoc test. The results were presented as mean ± standard deviation and statistical significance was considered for values of p < 0.05.

2.9 Research Ethical aspects

The protocol of animal use in the experiment was submitted and approved by the Comissão de Ética no Uso de Animais (CEUA, Animal Use Ethics Committee) of the University of Southern Santa Catarina under protocol number 19.007.4.01. IV. The Principles of Laboratory Animal Care, US National Institute of Health, NIH, were followed, as well as the Diretriz Brasileira para o Cuidado e a Utilização de Animais em Atividades de Ensino ou de Pesquisa Científica (DBCA, Brazilian Guideline for the Care and Use of Animals in Teaching or Scientific Research Activities) (2016), issued by the Conselho Nacional de Controle de Experimentação Animal (CONCEA, National Council for the Control of Animal Experimentation).

In our study, the effects of CSE on mice body weight, mesenteric fat weight, activity of complexes I and II of the mitochondrial respiratory chain in brain structures (hypothalamus, hippocampus, prefrontal cortex and striatum) and DNA damage in the cerebral cortex were evaluated in Swiss mice obesity-induced using a high-fat diet.

3.1 Body weight

After 7 weeks of obesity induction, before starting treatment with CSE, the mice body weight was evaluated; the obese mice group showed significant weight gain when compared to the control group, demonstrating that the animals that received a high-fat diet were obese. The results obtained after treatment with CSE did not demonstrate a statistically significant reversal of the weight of animals with obesity; however, those results did not indicate weight gain either (Fig. 2).

3.3 Mesenteric fat weight

Regarding the weight of mesenteric fat, animals that had received a high-fat diet (obese group) showed a significant increase in weight compared to animals that received a normal fat diet (control group); treatment with CSE was unable to reverse this increase (Fig. 3).

3.4 Mitochondrial respiratory chain complexes

Regarding the complexes evaluated in brain structures, complexes I and II showed a decrease in their activities in the obese group when compared to the control group, in all structures studied (Hypothalamus, prefrontal cortex, hippocampus and striatum) (Fig. 4 and Fig. 5). Treatment with CSE was able to reverse this damage in complex I in the hypothalamus (Fig. 4) and complex II in the prefrontal cortex (Fig. 5). However, even if this reversion was not observed in complex II in the hypothalamus, complex I in the cortex and the two complexes in the hippocampus and striatum (Fig. 4 and Fig. 5), the CSE was able to normalize the activity in these structures, since the treated obese group did not demonstrate statistical difference when compared with the control group.

3.5 Comet assay

In the comet assay, carried out in the cerebral cortex, diet-induced obese animals showed greater DNA damage compared to animals in the control group, and CSE was able to reverse this obesity damage (Fig. 6).

Currently, the compounds found in Cannabis sativa, such as THC, have been shown to have important effects on inflammation [41, 42]. As we already know, obesity is a low-grade chronic inflammation, responsible for causing countless damage to people’s health (de Bona Schraiber et al., 2019; Lin & Li, 2021; Usman & Volpi, 2018). Therefore, we ought to seek alternatives that can modulate and control the pathophysiology of obesity, as well as reverse its effects. For that reason, the present study evaluated the effects of THC-rich Cannabis sativa extract on obesity biochemical parameters.

To evaluate the effect of CSE on obesity, an obesity induction model was selected that uses mesenteric fat weight and body weight as an indicator. Initially, the body weight of the animals was reviewed, weight is indicative of fat accumulation in total adipose tissue, the growth of this indicator is associated with the onset of obesity [43]. Therefore, our findings demonstrated that, after the sixth week of a high-fat diet intake, the animals in the obesity group had greater body weight than those in the control group. Therefore, these data validate the obesity induction model used, as they corroborate the results by Kalupahana et al. [34] and Cintra et al [35]., where a similar model of high-fat diet-obesity induction in mice was used, for a period of 6 to 8 weeks, and the results demonstrated that there was an increase in body weight after consumption of this diet.

At the beginning of the eighth week of the experiment, the animals were treated orally with 1mL/kg/day CSE. The dosage and route of administration used in the study were based on the dosages prescribed and dispensed to patients from the partnering association which donated the CSE (unpublished data). The time set for the study is in accordance with Cluny et al. [45], who obtained satisfactory results with 3 weeks treatment.

Regarding body weight, CSE treatment has not been shown to reverse weight gain. Corroborating a study by Nguyen and collaborators [46], which reported that mice with high-fat diet-induced obesity, when treated with cannabis extract, did not experience significant changes in their body weight. Furthermore, our results demonstrate that the CSE treatment did not cause an increase in body weight as could be expected, based on a previous study by Zandani et al. [47], where a growth in body weight of animals fed a high-fat diet occurred even after treatment with cannabis extract. Corroborating the aforementioned, the study by Eitan et al [48]. also demonstrated that treatment with medicinal cannabis oils did not affect weight gain, caloric intake, or fat pad weight in mice fed a standard diet, although the size of adipocytes decreased in mice treated with THC-enriched extract.

We then evaluated mesenteric fat, as mouse mesenteric fat is considered the most analogous tissue to human intra-abdominal tissue [49], which excess is associated with the development of metabolic dysfunctions [50]. Therefore, our results indicate that the obesity group showed a significant difference compared to the control group. This result is in line with the study by de Mello et al. [51], where Swiss mice that received a high-fat diet demonstrated a significant difference in body weight, visceral fat and food consumption after six weeks of obesity induction when compared to the control group. Furthermore, the obesity-inducing model and the high-fat diet composition in our study was the same as that used in the work by de Mello et al. [51].

Regarding the group treated with CSE, there was no reduction in mesenteric fat compared to the untreated obese group. These findings contrast with data from a study by Cluny et al. [45], carried out in male C57BL obesity-induced mice with the use of a high-fat diet for 6 weeks and treated intraperitoneally with Δ-9-THC (2 mg/kg) or vehicle for 21 days and then THC (4 mg/kg) or vehicle for another 7 days. In addition to this, the study by Ramlugon et al. [52] demonstrated that a cannabis concentration of 1.25 mg/kg of body weight (in relation to THC content) was able to significantly reduce the average area of mesenteric fat adipocytes. Our study result demonstrated a reversal of the increase in body weight and fat mass in mice with obesity treated with THC 4 mg/kg/day in relation to mice without obesity. It is suggested that our findings can be explained by the low dosage we used, being equivalent to a dosage of 0.5mg/kg/day of THC and 0.04mg/kg/day of CBD.

In obesity, a greater concentration of CB1 receptors in the brain as well as in tissue growth can be found. THC can act as a functional antagonist, preventing the binding of endogenous agonists, acting as a modulator of food consumption [53]. Given this, it is possible to associate the results of Cluny et al. [45] with this effect. The present study, on the other hand, did not achieve the expected effect, and this result may be associated with the “entourage effect”, since the other compounds present in the CSE may have balanced the effect on CB1 [31, 32]. Furthermore, it is suggested that the effect on body weight may be dose dependent, since Cluny et al. [45] observed this effect only in the group that received a dosage of 4mg/kg/day. Furthermore, the intraperitoneal route may allow greater drug availability when compared to the oral route, the route used in our study [54].

There are reports that obesity is directly related to mitochondrial dysfunction making brain tissue susceptible to damage [55, 56]. Hence, the effects of obesity on complexes I and II of the mitochondrial respiratory chain, in the brain structures of the hypothalamus, hippocampus, striatum and prefrontal cortex, were also evaluated. Our findings demonstrate that there was a reduction in the activity of complexes I and II in all structures in the untreated obesity group. This reduction can be explained by the oxidative stress caused by the inflammatory process in obesity, and, as these are complexes responsible for capturing electrons from the Krebs cycle, for the formation of ATP, this damage can result in metabolic imbalance in all body systems and the worsening of already established oxidative stress [55]. The reduction in ATP formation in these structures is associated with functional cellular impairment, resulting in changes already observed in obesity, such as deficits in memory, cognition, learning, as well as imbalance in the reward system [12, 21, 55].

However, after the suggested treatment with CSE, normalization of complexes I and II activity occurred in all the structures assessed, since the treated group did not show a statistical difference when compared to the group that was not fed a high-fat diet. However, a statistically significant reversal was observed in the activity of complex I in the hypothalamus and of complex II in the prefrontal cortex, where an increase in the complexes’ activity was observed in these structures compared to the untreated obese group. Corroborating the findings of the present study, a review by Djeungoue-Petga and Hebert-Chatelain[57] brought together studies that showed that the CB1 receptor is present in large quantities in the hippocampus, hypothalamus and cerebral cortex, and its activation is associated with the modulation of activity of these structures.

The hypothalamus is a structure that is closely associated with the regulation of energy homeostasis, and just as inflammation has been reported in obesity, it has been demonstrated that there is a relationship between excess fat consumed in the diet and hypothalamic inflammation [58]. Furthermore, diets rich in fat end up compromising membrane fluidity, which can cause damage to mitochondrial membranes. However, our findings suggest that CSE treatment was able to suppress inflammation and reverse the likely hypothalamic damage caused by obesity, since Complex I is one of the main mediators of ROS [59]. On the other hand, the prefrontal cortex plays a fundamental role in cognition; however, these cognitive functions can be affected by high-fat diets and can bear negative impacts on glutamatergic neurotransmission and brain plasticity [60].

According to a study by Djeungoue-Petga and Hebert-Chatelain [57], there is expression of CB1 receptors in mitochondria at brain level, and their activation is associated with a reduction in cellular respiration and a decrease in ATP generation. These data suggest that isolated THC could promote responses contrary to those found in the present study, as seen in the work of Wolff et al. [61]. This last work demonstrated that THC could inhibit all complexes mitochondrial respiratory chain activity and, consequently, increasing the concentration of ROS in mice. In this connection it is suggested that the results obtained in the activity of complexes I and II of the respiratory chain are not linked to the action of THC on CB1 receptors.

Therefore, the reversal of the deficit detected in the respiratory chain complexes’ activity may be associated with the direct antioxidant effect of THC, enhanced by the presence of CBD, which in addition to its antioxidant potential, has a known anti-inflammatory effect, once again bringing the “entourage effect” into play. In this connection, both THC and CBD have aspects in their chemical structure that favor their interaction with ROS, directly neutralizing these substances. The antioxidant activities of these phytocannabinoids jointly or alone are so evident that they can be compared with the activity of vitamins E and C [29, 62].

The oxidative stress present in obesity is associated with the impairment of many cellular structures, including DNA [63, 64]. This information explains the findings of this study regarding an increased DNA damage in the cerebral cortex of the obesity group, since this is an inflammatory disorder associated with the generation of ROS [55, 65]. The cerebral cortex is a broad area of the brain, which can be associated with different functionalities and is closely linked to the general functioning of the organ [66, 67].

The cell DNA structural damage is linked to a range of metabolic impairments, such as difficulty in cell replication, imbalance in gene transcription associated with the formation of enzymes and hormones, for example [68]. In this connection, in our work we could observe the reversing effect of CSE on DNA damage, observed in the cerebral cortex of mice with obesity. There are no reports in the literature evaluating THC or CBD as substances directly protective against DNA damage. However, this finding is possibly linked to the antioxidant effect of THC associated with CBD [29]. These results may demonstrate the ability of THC to improve metabolic complications related to obesity when administered orally in increasing doses [48].

Therefore, the results found in this study demonstrate the potential of the CSE for the treatment of obesity, given its property of reversing damage to the respiratory chain and brain DNA. However, this study serves as a collaboration for new studies to be carried out, associating Cannabis sativa and THC with obesity

Obesity led to mitochondrial and DNA damage, and treatment with CSE proved to be effective in reversing these damages. It was observed that CSE treatment was able to reverse these damages in the mitochondrial respiratory chain complex I in the hypothalamus and complex II in the prefrontal cortex. Although no reversal was observed in the mitochondrial respiratory chain in hypothalamic complex II, cortical complex I, and hippocampal and striatal complexes, CSE was able to normalize activity in these structures, as the treated obese group showed no statistical difference when compared to the control group.

The results are due to the antioxidant action of THC, as well as the “entourage effect” promoted by all the compounds present in CSE. While the findings of this study are important for the scientific community, they deserve further studies that correlate THC and other components, as well as evaluate the effect of other concentrations of Cannabis sativa in the treatment of obesity.

Competing Interests

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Ethics approval

Funding

This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

Author Contribution

Ana Beatriz Costa: Conceptualization, Methodology, Formal analysis, Writing& Editing – Original Draft. Bruna Barros Fernandes: Conceptualization, Methodology, Writing – Review & Editing. Cristini da Rosa Turatti: Conceptualization, Methodology. Thalya Seifer Souza: Conceptualization, Methodology. Thais Medeiros de Jesus: Conceptualization, Methodology. Larissa Espindola da Silva: Conceptualization, Methodology. Mariana Pacheco de Oliveira: Conceptualization, Methodology. Mariella Reinol da Silva: Conceptualization, Methodology. Nicole Alessandra Engel: Conceptualization, Methodology. Daniéle Hendler Salla: Conceptualization, Methodology. Willian Sá Dias: Conceptualization, Methodology. Isabel Borges Becker: Conceptualization, Methodology. Adriani Paganini Damian: Conceptualization, Methodology. Larissa Barbosa: Conceptualization, Methodology. Luiza Martins Longaretti: Conceptualization, Methodology. Thais Ceresér Vilela: Conceptualization, Methodology. Renan Konig Leal: Writing – Review & Editing, Millena Fernandes: Writing – Review & Editing; Josiane Somariva Prophiro: Writing – Review & Editing, Vanessa Moraes de Andrade: Conceptualization, Methodology. Rafael Mariano De Bitencourt: Writing – Review & Editing, Supervision. Gislaine Tezza Rezin: Writing – Review & Editing, Supervision, Formal analysis.

Acknowledgement

We are grateful to the Anima Institute of Education, the National Council for Scientific and Technological Development (CNPQ), the Coordination for the Improvement of Higher Education Personnel (CAPES), and the Foundation for Research and Innovation in the State of Santa Catarina (FAPESC) for their support

Data Availability

All data generated or analyzed during this study are included in this paper and in their attached information files.

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Effects of THC-rich Cannabis sativa extract on Biochemical Parameters in Obesity

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Abstract

Figures

1. Introduction

2. Materials and Methods

2.1 Animals

2.2 Extract

2.3 Study design

2.4 Body weight and visceral fat

2.5 Tissue removal and storage

2.6 Mitochondrial respiratory chain enzymes activity analysis

2.7 DNA damage analysis

2.8 Statistical analysis

2.9 Research Ethical aspects

3. Results

3.1 Body weight

3.3 Mesenteric fat weight

3.4 Mitochondrial respiratory chain complexes

3.5 Comet assay

4. Discussion

5. Conclusion

Declarations

Competing Interests

Ethics approval

Funding

Author Contribution

Acknowledgement

Data Availability

References

Additional Declarations

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Version 1