Mosquitocidal and Anti-Inflammatory Properties of The Essential Oils Obtained from Monoecious, Male, and Female Inflorescences of Hemp (Cannabis sativa L.) and Their Encapsulation in Nanoemulsions.

Among the various innovative products obtainable from hemp (Cannabis sativa L.) waste biomass originating from different industrial processes, the essential oil (EO) deserves special attention in order to understand its possible application in different fields, such as cosmetics, pharmaceuticals, and botanical insecticides. For the purpose, in the present work, we studied the chemical composition of EOs obtained from different hemp varieties, namely Felina 32 and Carmagnola Selezionata (CS) using monoecious, male, and female inflorescences, and we evaluated their mosquitocidal activities on larvae and pupae of two main malaria vectors, Anopheles gambiae and An. stephensi. Then, in order to evaluate the safe use of hemp EOs for operators, the potential pro- or anti-inflammatory effect of hemp EOs together with their toxicological profile were determined on dermal fibroblasts and keratinocytes. Given the promising results obtained by insecticidal and anti-inflammatory studies, a preliminary evaluation of EOs encapsulation into nanoemulsions (NEs) has been performed with the aim to develop a formulation able to improve their poor physicochemical stability. Felina 32 and CS inflorescences provided EOs with an interesting chemical profile, with monoterpene and sesquiterpene hydrocarbons as the major components. This study highlighted the potential application of male inflorescences, which are usually discharged during hemp product processing. These EOs could be exploited as potential sustainable and eco-friendly insecticides, given their capability to be toxic against mosquitoes and the possibility to use them to prepare stable and safe formulations. The LC50 values found in this study (<80 ppm) are lower, on average, than those of many plant EOs, with the advantage of using an industrial waste product. From MTT assay and gene and protein expression analysis, EOs showed no cytotoxicity at the appropriate doses and exerted an anti-inflammatory effect on the human cell lines tested. These findings encourage further applied research on hemp EOs in order support their industrial exploitation.

A Clinical Evidence of a Correlation Between Insulin Resistance and the ALCAT Food Intolerance Test.

Insulin resistance (IR) is defined as the inability of a known quantity of exogenous or endogenous insulin to increase glucose uptake and utilization. Several mechanisms have been proposed as possible causes underlying the development of IR and the IR syndrome. IR occurs as part of a cluster of cardiovascular-metabolic abnormalities commonly referred to as "The Metabolic Syndrome." This may lead to the development of type 2 diabetes, accelerated atherosclerosis, hypertension, dysmenorrhea, hirsutism, and polycystic ovarian syndrome, depending on the genetic background of the individual developing IR. The aim of this study was to assess, in 123 female and 35 male (mean age, 42 y ± 10.3; range 19-75 y) volunteers) whether IR could be partly related to a dietary sugar intolerance and whether there could be a correlation between the ALCAT intolerance test and a mutation of the TCFTL2 gene (it promotes the trascription of the proglucagone and plays a key role in the development of the Langherans islands). Results evidenced that subjects with an intolerance to sugar, also showed a statistically significant complete or incomplete alteration of the TCFTL2 genetic test. Based upon these findings, our study demonstrated that there is a clinical correlation between the ALCAT food intolerance test and the IR. The positivity to the ALCAT test of one of the sugars tested (fructose, sugar cane, and sugar beet) indicates, in the majority of the subjects, the presence of a mutation of the gene TCF7L2 and could contribute to the prevention and treatment of the IR.

Effect of a probiotic intake on oxidant and antioxidant parameters in plasma of athletes during intense exercise training.

The aim of this study was to evaluate the effect of Lactobacillus rhamnosus IMC 501 and Lactobacillus paracasei IMC 502 on oxidative stress in athletes during a four-week period of intense physical activity. Two groups of twelve subjects each were selected for this analysis. The first group consumed a daily dose of a mixture of the two probiotic strains (1:1 L. rhamnosus IMC 501 and L. paracasei IMC 502; ~10(9) cells/day) for 4 weeks. The second group (control) did not consume any supplements during the 4 weeks. Blood samples collected immediately before and after the supplementation were analyzed, and plasma levels of reactive oxygen metabolites and biological antioxidant potential were determined. Faeces were also collected and analyzed before and at the end of the probiotic supplementation. Antioxidative activity and oxidative stress resistance of the two strains were determined in vitro. Results demonstrated that intense physical activity induced oxidative stress and that probiotic supplementation increased plasma antioxidant levels, thus neutralizing reactive oxygen species. The two strains, L. rhamnosus IMC 501(®) and L. paracasei IMC 502(®), exert strong antioxidant activity. Athletes and all those exposed to oxidative stress may benefit from the ability of these probiotics to increase antioxidant levels and neutralize the effects of reactive oxygen species.

Diaphragmatic breathing reduces exercise-induced oxidative stress.

Diaphragmatic breathing is relaxing and therapeutic, reduces stress, and is a fundamental procedure of Pranayama Yoga, Zen, transcendental meditation and other meditation practices. Analysis of oxidative stress levels in people who meditate indicated that meditation correlates with lower oxidative stress levels, lower cortisol levels and higher melatonin levels. It is known that cortisol inhibits enzymes responsible for the antioxidant activity of cells and that melatonin is a strong antioxidant; therefore, in this study, we investigated the effects of diaphragmatic breathing on exercise-induced oxidative stress and the putative role of cortisol and melatonin hormones in this stress pathway. We monitored 16 athletes during an exhaustive training session. After the exercise, athletes were divided in two equivalent groups of eight subjects. Subjects of the studied group spent 1 h relaxing performing diaphragmatic breathing and concentrating on their breath in a quiet place. The other eight subjects, representing the control group, spent the same time sitting in an equivalent quite place. Results demonstrate that relaxation induced by diaphragmatic breathing increases the antioxidant defense status in athletes after exhaustive exercise. These effects correlate with the concomitant decrease in cortisol and the increase in melatonin. The consequence is a lower level of oxidative stress, which suggests that an appropriate diaphragmatic breathing could protect athletes from long-term adverse effects of free radicals.

Effects of Two Months of Very Low Carbohydrate Ketogenic Diet on Body Composition, Muscle Strength, Muscle Area, and Blood Parameters in Competitive Natural Body Builders.

Background: Ketogenic diet (KD) is a nutritional approach that restricts daily carbohydrates, replacing most of the reduced energy with fat, while maintaining an adequate quantity of protein. Despite the widespread use of KD in weight loss in athletes, there are still many concerns about its use in sports requiring muscle mass accrual. Thus, the present study sought to investigate the influence of a KD in competitive natural body builders. Methods: Nineteen volunteers (27.4 ± 10.5 years) were randomly assigned to ketogenic diet (KD) or to a western diet (WD). Body composition, muscle strength and basal metabolic rate were measured before and after two months of intervention. Standard blood biochemistry, testosterone, IGF-1, brain-derived neurotrophic factor (BDNF) and inflammatory cytokines (IL6, IL1ß, TNFα) were also measured. Results: Body fat significantly decreased in KD (p = 0.030); whilst lean mass increased significantly only in WD (p < 0.001). Maximal strength increased similarly in both groups. KD showed a significant decrease of blood triglycerides (p < 0.001), glucose (p = 0.001), insulin (p < 0.001) and inflammatory cytokines compared to WD whilst BDNF increased in both groups with significant greater changes in KD (p < 0.001). Conclusions: KD may be used during body building preparation for health and leaning purposes but with the caution that hypertrophic muscle response could be blunted.

Inhibition of adrenocortical carcinoma by diphtheria toxin mutant CRM197.

BACKGROUND: In this study, we investigated the effect of CRM197 treatment in human adrenocortical carcinoma (AC) implanted in nude mice. CRM197 is a non-toxic mutant of diphtheria toxin that binds heparin-binding epidermal growth factor-like growth factor (HB-EGF) which is implicated in the proliferative activity of several tumor cells. METHODS: HB-EGF expression in AC cells was evaluated by reverse transcription PCR and Western blot. AC tumors were implanted in nude mice and then treated with CRM197. Effects of treatment on angiogenesis and apoptosis were investigated by immunohistochemistry and Western blot. The effects on cell invasion and migration were investigated with a matrigel invasion assay. RESULTS: We demonstrated that human AC cells express HB-EGF. A treatment with CRM197 blocked growth, reduced angiogenesis and induced apoptosis in AC tumors implanted in nude mice. CRM197 also inhibited invasion and migration of these tumor cells. CONCLUSIONS: These data support the evidence for anticancer properties of CRM197 in AC tumors.

Ketogenic Diet and Skeletal Muscle Hypertrophy: A Frenemy Relationship?

Ketogenic diet (KD) is a nutritional regimen characterized by a high-fat and an adequate protein content and a very low carbohydrate level (less than 20 g per day or 5% of total daily energy intake). The insufficient level of carbohydrates forces the body to primarily use fat instead of sugar as a fuel source. Due to its characteristic, KD has often been used to treat metabolic disorders, obesity, cardiovascular disease, and type 2 diabetes. Skeletal muscle constitutes 40% of total body mass and is one of the major sites of glucose disposal. KD is a well-defined approach to induce weight loss, with its role in muscle adaptation and muscle hypertrophy less understood. Considering this lack of knowledge, the aim of this review was to examine the scientific evidence about the effects of KD on muscle hypertrophy. We first described the mechanisms of muscle hypertrophy per se, and secondly, we discussed the characteristics and the metabolic function of KD. Ultimately, we provided the potential mechanism that could explain the influence of KD on skeletal muscle hypertrophy.

Mebendazole inhibits growth of human adrenocortical carcinoma cell lines implanted in nude mice.

Adrenocortical carcinoma is a rare tumor of the adrenal gland which requires new therapeutic approaches as its early diagnosis is difficult and prognosis poor despite therapies used. Recently, mebendazole has been proved to be effective against different cancers. The aim of our study was to evaluate whether mebendazole may result therapeutically useful in the treatment of human adrenocortical carcinoma. We analyzed the effect of mebendazole on human adrenocortical carcinoma cells in vitro and after implantation in nude mice. In order to clarify mechanisms of mebendazole action, metastases formation, apoptosis and angiogenesis were also investigated. Mebendazole significantly inhibited cancer cells growth, both in vitro and in vivo, the effects being due to the induction of apoptosis. Moreover, mebendazole inhibited invasion and migration of cancer cells in vitro, and metastases formation in vivo. Overall, these data suggest that treatment with mebendazole, also in combination with standard therapies, could provide a new protocol for the inhibition of adrenocortical carcinoma growth.

Body composition obtained from the body mass index: an Italian study.

BACKGROUND: Since obesity and related diseases are now considered epidemic, new and more accurate formulas for epidemiological studies are of interest to the scientific community. Several equations have been proposed to estimate the body composition simply from anthropometric measurements. However, with time, the body composition of the populations studied changes in relation to their food habits and lifestyle, and, therefore, the equations must be regularly updated and corrected. AIM OF THE STUDY: The aim of the study was to develop new equations to determine the body composition among the Italian population using the body mass index and independently by variables such as age and body structure. METHODS: Bioelectrical impedance and anthropometric analysis of 764 Italian Caucasian subjects (342 females and 422 males), 11 to 80 years of age, were analysed. Females and males were analysed separately. Multiple regression analyses were performed in order to estimate the body composition of the subjects. The estimated masses were then compared with the measured masses using Bland and Altman plots. We also calculated the differences between the estimated and measured masses, reported as % of the body weight, for the 95, 85 and 75 degrees percentile of the female and male groups. Finally we compared our formulas with the Watson equations, which are used to estimate the total body water. RESULTS: All body masses estimated were positively correlated to the measured values. Moreover, at any percentile analysed, our formulas resulted more precise than the Watson formula. Equations: Females: FM = 1.9337 BMI - 26.422; FFM = BW - FM; BCM = 0.3655 FFM + 4.865; TBW = 0.5863 FFM + 7.1732; Males: FM = 1.407 BMI - 21.389; FFM = BW - FM; BCM = 0.4485 FFM + 3.3534; TBW = 0.6997 + 1.4567. CONCLUSIONS: Although an inevitable inaccuracy must be expected in epidemiological studies, our equations are adequate to analyze the body composition state and changes occurring among the Italian population by simply considering weight and height.

Effects of losartan and irbesartan administration on brain angiotensinogen mRNA levels.

Losartan, 2-n-butyl-4-chloro-5-hydroxymethyl-1-[(2'(1H-tetrazol-5-yl)-biphenil-4-yl)methyl]imidazole, and Irbesartan, 2-n-butyl-3-[(2'-(1H-tetrazol-5-yl)-biphenyl-4-yl)methyl]-1,3-diaza-spiro[4,4]non-1-en-4-one, are two angiotensin AT1 receptor antagonists largely used in human health care as antihypertensive agents. Their ability to cross the blood-brain barrier and to influence the central renin-angiotensin system are widely investigated, but how this brain system responds to the subchronic and chronic block of the angiotensin AT1 receptor is still unknown. Normotensive rats were intragastrically implanted for 7- and 30-day administration, with a dose of 3 and 30 mg/kg body weight. Treatments were shown to influence, in a dose-, time- and brain-area-dependent manner, angiotensinogen mRNA levels in scanned areas. This study showed a general up-regulation of angiotensinogen mRNA expression after 7 days and a widespread down-regulation or basal level of expression after a 30-day administration of two angiotensin AT1 receptor antagonists.

Preprotachykinin A gene expression after administration of 3,4-methylene dioxymethamphetamine (Ecstasy).

This study tested the effects of 8 days of subchronic administration of 3,4-methylene dioxymethamphetamine (MDMA) (5 mg/kg b.w.) on preprotachykinin A mRNA levels in discrete rat brain regions. In situ hybridization examined preprotachykinin A mRNA levels in the core and shell of the nucleus accumbens, the islands of Calleja, the olfactory tubercle, the dorsal and ventral caudate-putamen, the bed nucleus of the stria terminalis, the medial preoptic area, the medial habenular nucleus and in the postero-dorsal part of the medial amygdala. Higher levels of preprotachykinin A mRNA were found in the core and shell of the nucleus accumbens, in the islands of calleja, in the olfactory tubercle, in the bed nucleus of the stria terminalis, in the medial habenular nucleus and the postero-dorsal part of the medial amygdala, compared to control animals. Conversely, increased preprotachykinin A mRNA levels were observed in the dorsal and ventral caudate-putamen in MDMA treated when compared to control rats. In the social memory test, MDMA significantly impaired rats' short-term working memory. These results show that chronic exposure to MDMA strongly affects preprotachykinin A mRNA levels in discrete rat brain regions. These changes occur in experimental conditions in which working memory is markedly reduced, suggesting that changes in gene expression of tachykinin mechanisms may contribute to the effects of MDMA on memory function.

Memory impairment following combined exposure to delta(9)-tetrahydrocannabinol and ethanol in rats.

Cannabis derivatives and alcohol are widely co-abused, particularly among adolescents. Since both ethanol and cannabinoids are known to impair learning and memory, the present study investigated in rats the effects of combined exposure to ethanol and delta(9)-tetrahydrocannabinol (THC) in a memory task, the object recognition test. The results of the present study provide evidence that ethanol, voluntarily ingested in alcohol-preferring rats, and THC, given by intraperitoneal injection, have a synergic action to impair object recognition, when a 15-min interval was adopted between the sample phase and the choice phase of the test. Neither voluntary ethanol ingestion nor 2 or 5 mg/kg of THC were able per se to modify object recognition in these experimental conditions, but when voluntary ethanol ingestion was combined with administration of these doses of THC object recognition was markedly impaired. THC impaired object recognition only at the dose of 10 mg/kg, when its administration was not combined with that of ethanol. The selective cannabinoid CB(1) receptor antagonist SR 141716A (N-(piperidin-1-yl)-5-(4-chlorophenyl)-1(2, 4-dichloro-phenyl)-4-methyl-1H-pyrazole carboxamide.HCl) at the dose of 1 mg/kg reversed the amnesic effect of THC, 10 mg/kg, suggesting that the effect is mediated by this receptor subtype. The synergism of ethanol and THC was not detected when an inter-trial interval of 1 min was adopted. The present findings are in keeping with the notion that Cannabis derivatives impair memory processes and provide evidence for a synergic action of THC and ethanol, thus emphasizing the risks consequent to their co-administration.

Diaphragmatic breathing reduces postprandial oxidative stress.

OBJECTIVES: A number of studies suggest that postprandial hyperglycemia produces oxidative stress, leading to complications associated with diabetes. However, hyperglycemia-induced oxidative stress may affect groups of people other than diabetics, such as smokers and athletes with specific diet plans. Based on previous reports that seated breathing meditation reduces hyperglycemia, the present study was designed to determine the effects of diaphragmatic breathing on postprandial plasma glycemia, insulin, oxidative stress, and antioxidant levels in athletes with normal glucose metabolism. DESIGN: Data collected before and after consumption of a 900-calorie breakfast composed of 80% carbohydrates, 10% proteins, and 10% lipids were analyzed. Ten (10) minutes after the meal, 8 subjects spent 40 minutes performing diaphragmatic breathing in a quiet place. The other 8 subjects, representing the control group, spent the same time sitting in an equivalent quiet place reading a magazine. SUBJECTS: Data from 16 amateur male cyclists age 30.12±4.9 years (±SD) were analyzed. Their mean height and weight were 177.81±5.3 cm and 71.40±5.2 kg, respectively. All subjects underwent a physical examination and were determined to be in good health. OUTCOME MEASURES: Blood samples were collected immediately before the meal as well as 1 hour and 2 hours after the meal, and plasma levels of glucose, insulin, reactive oxygen metabolites, and biologic antioxidant potential were determined. Heart rate was also recorded. RESULTS: Results show that in normal subjects, acute hyperglycemia induces free-radical production while reducing the antioxidant levels (p<0.05). Diaphragmatic breathing reduces heart rates (p<0.01), increases insulin (p<0.05), reduces glycemia (p<0.01), and reduces free-radical production as indicated by the higher antioxidants levels (p<0.05). CONCLUSIONS: Diaphragmatic breathing, likely through the activation of the parasympathetic nervous system, increases insulin, reduces glycemia, and reduces reactive oxygen species production.

Doxazosin-related alpha1-adrenoceptor antagonists with prostate antitumor activity.

Doxazosin analogues 1-3 and 1a were synthesized and investigated at alpha1-adrenoceptors and PC-3, DU-145, and LNCaP human prostate cancer cells. Compound 1 (cyclodoxazosin) was a potent alpha(1B)-adrenoceptor antagonist displaying antiproliferative activity higher than that of doxazosin in cancer cells in vitro and in vivo, respectively. Because of its antitumor efficacy at low concentrations, lower apoptotic activity in NHDF vs tumor cells, and antiangiogenetic effect, 1 showed a better therapeutic profile relative to doxazosin.

Neonatal capsaicin treatment affects rat thymocyte proliferation and cell death by modulating substance P and neurokinin-1 receptor expression.

Herein we provide evidence that substance P (SP) and its neurokinin-1 receptor (NK-1R) expressed on thymocytes counteract thymus depletion induced by neonatal capsaicin (CPS) treatment by affecting thymocyte proliferation and apoptotic death. SP administration reversed the CPS-mediated inhibitory effects on the total thymocyte number and subset distribution, namely CD4+ and CD4- CD8- cells, through its interaction with NK-1R as shown by concomitant NK-1R (SR140333) antagonist administration. SP-induced enhancement of thymus cellularity parallels its ability of inhibiting the thymocyte apoptotic program. Indeed, exogenously administered SP completely nullified CPS-induced apoptosis, and SR140333 abrogated the SP-mediated protective effect. SP administration also stimulated concanavalin A (Con A)-induced thymocyte proliferation of CPS-treated rats, completely reversing the CPS-induced inhibition. The SP-mediated stimulation of Con A-induced thymocyte proliferation was NK-1R dependent as shown by concomitant administration of SP and SR140333 to CPS-treated rats. Our results also demonstrate that CPS treatment induces a marked decrease of thymocyte PPT-A mRNA level and endogenous SP content as evaluated by quantitative RT-PCR, in situ hybridization and cytofluorimetric analysis. By contrast, NK-1R mRNA levels were increased in thymocytes from CPS-treated rats. Exogenous SP administration augmented PPT-A, SP and NK-1R thymocyte expression in CPS-treated rats, and this enhancement was antagonized by SR140333 administration. Overall, our results strongly suggest that the immunomodulatory effects of neonatal CPS treatment on rat thymocyte functions are dependent on vanilloid-mediated regulation of SP and NK-1R functional expression by neuronal and immune cells.

Expression of substance P and its neurokinin-1 receptor on thymocytes: functional relevance in the regulation of thymocyte apoptosis and proliferation.

Herein, we provide evidence of the expression and function of substance P (SP) and neurokinin-1 receptor (NK-1R) in the rat thymus. In situ hybridization evidenced NK-1R mRNA mainly in the thymic medulla, and Northern blot analysis of mRNA from FACS-sorted thymocytes identified NK-1R on CD4+, CD8+ and double-positive subpopulations. With flow cytometry, it could be seen that NK-1R was expressed on the majority of CD5+ thymocytes, and it was identified by Western blot analysis as two bands migrating at 44 and 54 kD. SP administration rescues thymocytes from spontaneous and NK-1R antagonist (SR140333)-induced apoptosis and stimulates concanavalin A (ConA)-induced thymocyte proliferation, CD25 expression and IL-2 production, whereas SR140333 exerts inhibitory effects on these functions. We also demonstrated the expression of mRNA for the SP precursor preprotachykinin-A in the thymic medulla and purified CD5+ thymocytes. SP protein was detected on 40% of CD5+ thymocytes and identified as a band of 1.3 kD by Western blot analysis. Finally, thymocytes spontaneously released SP, which was increased upon ConA or CD3 stimulation.