Organ Weight Changes Research Articles

In Vitro and In Vivo Toxicological Evaluation of Avicennia africana P: Beauv. (Avicenniaceae) Leaf Extract in a Rat Model.

Avicennia africana is an important ethnomedicinal plant that has long been used to treat malaria and several other diseases. Despite the plant's antimalarial and other therapeutic properties, there is limited evidence-based data on its potential toxicity. Hence, the purpose of the current study was to assess the safety of A. africana leaf ethanolic extract (AAE). The study was designed to ascertain the cytotoxic effects of the crude extract on red blood cells (RBCs) as well as the acute and subacute toxicity in Wistar albino rats in accordance with Organization for Economic Co-operation and Development (OECD) guidelines "Test No. 423" and CPMW/SWP/1042/99. The pulverized, shade-dried plant leaves were sequentially macerated with 70% ethanol to obtain the crude extract (AAE). The extract's cytotoxic activity (CC50) against the uninfected human red blood cells (RBCs) was determined using the 3-(4,5-Dimethylythiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. For the acute toxicity studies, the rats (male and female) were divided randomly into six groups of five rats (n = 5) and dosed orally once with the following dose levels: 100, 300, 1000, 3000, and 5000 mgkg-1, p.o. of the extracted AAE, with the control group receiving only the vehicle. In the repeated dose toxicity studies, the rats (both sexes) were orally administered daily with AAE at 100, 300, and 1000 mgkg-1 for 14 days. Rat body weights were measured, and blood samples were tested for haematological and biochemical markers. Internal organs like the heart, kidney, liver, and spleen were collected, inspected, and weighed, and histological examinations were performed. The median lethal dose (LD50) value is greater than 5000 mgkg-1 body weight, with no significant change in bodyweight or relative organ weight (ROWs) of the extract-treated groups or control group. The extract showed greater cytotoxicity activity (CC50), which was >100 μg/mL, compared to the reference drug (artesunate).The dosage groups of 100 and 300 mgkg-1bwt had neutrophilia and lymphocytopenia (p < 0.05). However, changes in these haematological parameters may not be dose dependent and could be stress related. All the serum biochemical markers studied in rats given AAE did not show any significant change (p > 0.05). Histopathological examination of internal organs of AAE-treated rats did not show any significant abnormalities resulting from the extract treatment compared to the control group. Based on the findings in the present study, the LD50 value of AAE was found to exceed 5000 mgkg-1 in the acute toxicity test, while the no observed adverse effect level (NOAEL) in rats was 1000 mgkg-1 p.o. In the sub-acute toxicity tests. Histopathological analysis revealed no morphological abnormalities in the vital organs.

Long-term evaluation of safety and biological effects of Korean Red Ginseng (Panax Ginseng): a long-term in vivo study

BackgroundAlthough Korean Red Ginseng (KRG) is safe, this finding was only evaluated in 3-mo-long studies. Its safety was verified through a 6-mo KRG administration clinical study, but long-term studies beyond 6 mo are insufficient. This study investigated the safety and efficacy of 12-mo KRG administration.MethodsIn this study, 300 mg/kg of KRG was administered to male and female Sprague Dawley rats for 4, 8, and 12 mo to evaluate its efficacy and safety. Clinical signs, including pathological examination and haematological analyses, were observed. Flow cytometric analyses were utilised to analyse spleen and thymus immune cell counts after 12 mo. Proteomic analysis of the sera was performed using a nanospray-interfaced mass spectrometer with an 11-plex Tandem Mass Tag (TMT) labelling system. Bioinformatic analysis was then performed using Ingenuity Pathway Analysis and PANTHER. Data are available via ProteomeXchange with identifier PXD032036.ResultsNo significant body and organ weight changes were observed, and haematological and serum biochemical analyses did not show clinical significance. The effectiveness of long-term KRG administration was confirmed through increased immune cell distribution and activity. Changes in proteins correlated with viral infection reduction were confirmed through proteomic analysis.ConclusionThe results suggested that 12-mo KRG intake is safe, improves immune system activity, and reduces viral infections with no significant changes in toxicological aspects.

Safety assessment of oil extracted from lacquer (Toxicodendron vernicifluum (Stokes) F.A. Barkley) seed: acute and subchronic toxicity studies in rats

Ethnopharmacological relevanceToxicodendron vernicifluum (Stokes) F.A. Barkley (RVS) is an economic tree species and widely distributed in East Asia. Wood parts and raw lacquers of RVS have been used in coatings, herbal medicines or food supplements, and the leaves, flowers, roots, and fruits of RVS are also widely used in medicine traditionally. Lacquer seed oil (LSO) has potential health benefits and has not previously been evaluated for safety. Aim of the studyThe aim of the present study was to investigate the toxicological potential of LSO by acute and subchronic toxicity tests. Materials and methodsThe characterization of fatty acids of the LSO was carried out by gas chromatography. In the acute toxicity study, LSO was administered at single doses of 5000 or 10000 mg/kg by oral gavage. The subchronic toxicity study was conducted by daily oral administration of LSO at doses of 1250, 2500 and 5000 mg/kg/day for 30 consecutive days. The animals were evaluated for clinical observations, body weight, organ weight, feed consumption, biochemical and hematological parameters, and liver, lung, and kidney histology. ResultsThere were no mortality and toxic changes were observed in acute toxicity study. The results of subchronic toxicity showed no toxicologically significant changes in clinical observations, body weight, organ weight, biochemical or hematological parameters. Histopathologic results indicated slight hepatic steatosis and inflammatory infiltration in the rats of 5000 mg/kg/day LSO treated group. However, the histopathologic observation was not confirmed by hepatic biochemical analysis. ConclusionsThese results suggested that the LD50 of LSO is over 10000 mg/kg and LSO is non-toxic for SD rats in acute toxicity study. The no observed adverse effect level (NOAEL) of LSO in rats is considered to be 5000 mg/kg/day, and liver is the potential target organ of LSO for 30-day subchronic toxicity study.

Acute and Subacute Toxicity Studies of Erodium guttatum Extracts by Oral Administration in Rodents.

The present study aimed to evaluate the acute and subacute toxicity profiles of Erodium guttatum extracts in mice using the methods described in the guidelines of the OECD. In the acute toxicity study, the LD50 value was greater than 2000 mg/kg. The subacute toxicity study of E. guttatum extracts showed no significant changes in body or organ weights. The administration of E. guttatum extracts to mice at a dose of 200 mg/kg led to an increase in white blood cells, platelets and hemoglobin. Moreover, the aqueous extract of E. guttatum only decreased liver aspartate aminotransferase (ASAT) levels at a dose of 200 mg/kg, and creatinine and urea levels did not show any significant alterations compared to the control group. Our results showed that the extracts of E. guttatum caused a slight increase in alanine aminotransferase (ALAT) and triglycerides. The histological study showed that mice treated with E. guttatum extracts experienced some histopathological changes in the liver, particularly with the methanolic extract, and slight changes in the kidneys and pancreas. Regarding the renal profile, no toxicity was observed. These results provide basic information on the toxicological profile of E. guttatum used in traditional medicine.

The Acute and Short-Term Inhalation of Carbon Nanofiber in Sprague-Dawley Rats.

The inhalation toxicity of carbon nanofibers (CNFs) is not clearly known due to relatively few related studies reported. An acute inhalation study and short-term inhalation study (5 days) were therefore conducted using Sprague-Dawley rats. In the acute inhalation study, the rats were grouped and exposed to a fresh air control or to low (0.238 ± 0.197), moderate (1.935 ± 0.159), or high (24.696 ± 6.336 mg/m3) CNF concentrations for 6 h and thereafter sacrificed at 14 days. For the short-term inhalation study, the rats were grouped and exposed to a fresh air control or low (0.593 ± 0.019), moderate (2.487 ± 0.213), or high (10.345 ± 0.541 mg/m3) CNF concentrations for 6 h/day for 5 days and sacrificed at 1, 3, and 21 days post-exposure. No mortality was observed in the acute inhalation study. Thus, the CNF LC50 was higher than 25 mg/m3. No significant body or organ weight changes were noted during the 5 days short-term inhalation study or during the post-exposure period. No significant effects of toxicological importance were observed in the hematological, blood biochemical, and coagulation tests. In addition, the bronchoalveolar lavage (BAL) fluid cell differential counts and BAL inflammatory markers showed no CNF-exposure-relevant changes. The histopathological examination also found no CNF-exposure-relevant histopathological lesions. Thus, neither acute nor 5 days inhalation exposure to CNFs induced any noticeable toxicological responses.

Cytotoxicity and acute toxicity evaluation of hydrogen-rich Gynostemma pentaphyllum Makino distillate

Hydrogen-rich Gynostemma pentaphyllum Makino distillate (HRGD) consists of Gynostemma pentaphyllum Makino steam distillate with hydrogen gas. Although both G. pentaphyllum Makino and hydrogen-rich water are well known for their biological and medical benefits, there is a lack of information on their safety and toxicity in vivo acute oral toxicity test and in vitro cytotoxicity method. The current study aimed to assess the cytotoxicity and acute oral toxicity of HRGD as a part of a safety evaluation using rat and human cell models. HRGD was administered orally once by gavage to male and female Sprague-Dawley rats at doses of 0, 2500, and 5000 mg/kg. Cytotoxicity assay was conducted in vitro at various concentrations in 10 different human normal and cancer cell lines; TK6 (human normal lymphomablastoid cells), Chang (human hepatic cells), 16HBE14o- (human bronchial epithelial cells), URotsa (human urothelium cells), MCF (human breast cancer cells), Hela (human cervical cancer cells), A375 (human malignant melanoma cells), HCT116 (human colon cancer cells), HepG2 (human liver cancer cells) and A549 (human non-small cell lung adenocarcinoma cells). From a 14-day study in rats, we observed no compound-related changes in mortality, clinical signs, body weight, food/water consumption, organ weight and gross pathology in all dose group. The result of in vivo acute toxicity shows that no observed adverse effect level of HRGD was below 5000 mg/kg for both sexes of rats, and the minimal lethal dose was considered to be more than 5000 mg/kg. HRGD also had no in vitro cytotoxicity against all tested cells. The present study data indicated that HRGD may contain bioactive compounds of potential therapeutic significance that are relatively safe from toxic effects.

Development, Validation, and Application of a Human Reproductive Toxicity Prediction Model Based on Adverse Outcome Pathway.

A growing number of environmental contaminants have been proved to have reproductive toxicity to males and females. However, the unclear toxicological mechanism of reproductive toxicants limits the development of virtual screening methods. By consolidating androgen (AR)-/estrogen receptors (ERs)-mediated adverse outcome pathways (AOPs) with more than 8000 chemical substances, we uncovered relationships between chemical features, a series of pathway-related effects, and reproductive apical outcomes─changes in sex organ weights. An AOP-based computational model named RepTox was developed and evaluated to predict and characterize chemicals' reproductive toxicity for males and females. Results showed that RepTox has three outstanding advantages. (I) Compared with the traditional models (37 and 81% accuracy, respectively), AOP significantly improved the predictive robustness of RepTox (96.3% accuracy). (II) Compared with the application domain (AD) of models based on small in vivo datasets, AOP expanded the ADs of RepTox by 1.65-fold for male and 3.77-fold for female, respectively. (III) RepTox implied that hydrophobicity, cyclopentanol substructure, and several topological indices (e.g., hydrogen-bond acceptors) were important, unbiased features associated with reproductive toxicants. Finally, RepTox was applied to the inventory of existing chemical substances of China and identified 2100 and 7281 potential toxicants to the male and female reproductive systems, respectively.

Fetal programming in sheep: Effects on pre- and postnatal organs and glands development in lambs

The present systematic review and meta-analysis aim to summarize the effects of maternal undernutrition or overnutrition during pregnancy on the absolute weight and relative weight of the organs (liver, kidneys, heart, spleen, and lung) and glands (adrenal, pancreas, and thyroid) measured during gestation, birth and the postnatal period in lambs. After completing the search, selection, and data extraction steps, the measure of effect was generated by the individual comparison of each variable response compared with the average of the control and treated group (undernutrition or overnutrition) using the DerSimonian and Laird method for random effects. The liver was the organ most affected by maternal undernutrition, as the absolute weight of the liver was reduced during pregnancy, birth, and the postnatal period. The extent of this effect is related to the duration of the intervention. Reductions in the absolute fetal weight of the lungs and spleen have also been observed. No change in organs weight were observed when the results were expressed as relative weight. For overnutrition, the fetal weight of the liver was reduced to both absolute and relative values. In contrast, the relative weight of the kidneys has been increased. For the glands analyzed, no changes in weight were observed in either scenario (absolute or relative weight). Thus, the organs are more likely to suffer weight changes, especially during pregnancy, as a result of maternal nutrition. However, this change in organ weight seems to be closely related to the reduction in body weight of the progeny as a whole.

Abstract LB203: Assessment of the safety, pharmacokinetics, and pharmacodynamics of a first-in-class cancer drug candidate E-602, a sialoglycan degrader, in non-human primates

Abstract E-602 is a novel, first-in-class, engineered human sialidase (Neu2) Fc fusion developed for cancer treatment by enhancing antitumor immunity through desialylating immunosuppressive sialoglycans. We have previously shown that E-602 cleaves off terminal sialic acids from surface sialoglycans on tumor and immune cells, restores the immune function of exhausted-like T cells, and enhances dendritic cell priming and activation of naïve T cells. E-602 has also demonstrated antitumor activity as a single agent in multiple syngeneic mouse tumor models. To support clinical development, we conducted a Good Laboratory Practice one-month repeat-dose toxicity study to evaluate E-602’s pharmacokinetic (PK) parameters, pharmacodynamic (PD) effects, and safety profiles in cynomolgus monkeys via weekly intravenous infusion of E-602 at 5, 30, 100, and 200 mg/kg for five doses. E-602 showed an apparent half-life (t1/2) of 4-6 hours and the terminal t1/2 of 10-30 hours with dose linearity from 30 mg/kg to 200 mg/kg. E-602 produced dose-dependent PD effects of desialylation of immune cells, such as T cells, monocytes, and dendritic cells, as monitored by flow cytometry. Despite the short half-life of E-602, the PD effect of immune cell desialylation sustained up to seven days for T cells, consistent with E-602’s enzymatic mechanism of action. Generally, E-602 was well tolerated up to 100 mg/kg with no dose-limiting adverse E-602-related changes in body weight, ophthalmology, neurological/musculoskeletal assessments, electrocardiogram parameters, respiratory rate, coagulation, urinalysis, organ weights, and histopathology. Animals in the 200 mg/kg dose group survived but showed isolated adverse histopathological changes with an uncertain relationship to E-602 administration. Therefore, the no-observed-adverse-effect-level (NOAEL) of E-602 was determined as 100 mg/kg in cynomolgus monkeys. In conclusion, the PK and PD data have supported the selection of cynomolgus monkeys as the toxicity species to evaluate E-602’s safety profile. E-602 has demonstrated sustained PD effects of immune cell desialylation up to seven days postdose and a wide safety margin with NOAEL of 100 mg/kg in cynomolgus monkeys. Citation Format: Lizhi Cao, Jenny Che, Ali Chesney, Rakesh Dixit, Nancy Zheng, Nicole Zane, James Broderick, Li Peng. Assessment of the safety, pharmacokinetics, and pharmacodynamics of a first-in-class cancer drug candidate E-602, a sialoglycan degrader, in non-human primates [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB203.

Repository Describing the Anatomical, Physiological, and Biological Changes in an Obese Population to Inform Physiologically Based Pharmacokinetic Models.

BackgroundObesity is associated with physiological changes that can affect drug pharmacokinetics. Obese individuals are underrepresented in clinical trials, leading to a lack of evidence-based dosing recommendations for many drugs. Physiologically based pharmacokinetic (PBPK) modelling can overcome this limitation but necessitates a detailed description of the population characteristics under investigation.ObjectiveThe purpose of this study was to develop and verify a repository of the current anatomical, physiological, and biological data of obese individuals, including population variability, to inform a PBPK framework.MethodsA systematic literature search was performed to collate anatomical, physiological, and biological parameters for obese individuals. Multiple regression analyses were used to derive mathematical equations describing the continuous effect of body mass index (BMI) within the range 18.5–60 kg/m2 on system parameters.ResultsIn total, 209 studies were included in the database. The literature reported mostly BMI-related changes in organ weight, whereas data on blood flow and biological parameters (i.e. enzyme abundance) were sparse, and hence physiologically plausible assumptions were made when needed. The developed obese population was implemented in Matlab® and the predicted system parameters obtained from 1000 virtual individuals were in agreement with observed data from an independent validation obese population. Our analysis indicates that a threefold increase in BMI, from 20 to 60 kg/m2, leads to an increase in cardiac output (50%), liver weight (100%), kidney weight (60%), both the kidney and liver absolute blood flows (50%), and in total adipose blood flow (160%).ConclusionThe developed repository provides an updated description of a population with a BMI from 18.5 to 60 kg/m2 using continuous physiological changes and their variability for each system parameter. It is a tool that can be implemented in PBPK models to simulate drug pharmacokinetics in obese individuals.

Time course of pulmonary inflammation and trace element biodistribution during and after sub-acute inhalation exposure to copper oxide nanoparticles in a murine model

BackgroundIt has been shown that copper oxide nanoparticles (CuO NPs) induce pulmonary toxicity after acute or sub-acute inhalation exposures. However, little is known about the biodistribution and elimination kinetics of inhaled CuO NPs from the respiratory tract. The purposes of this study were to observe the kinetics of pulmonary inflammation during and after CuO NP sub-acute inhalation exposure and to investigate copper (Cu) biodistribution and clearance rate from the exposure site and homeostasis of selected trace elements in secondary organs of BALB/c mice.ResultsSub-acute inhalation exposure to CuO NPs led to pulmonary inflammation represented by increases in lactate dehydrogenase, total cell counts, neutrophils, macrophages, inflammatory cytokines, iron levels in bronchoalveolar lavage (BAL) fluid, and lung weight changes. Dosimetry analysis in lung tissues and BAL fluid showed Cu concentration increased steadily during exposure and gradually declined after exposure. Cu elimination from the lung showed first-order kinetics with a half-life of 6.5 days. Total Cu levels were significantly increased in whole blood and heart indicating that inhaled Cu could be translocated into the bloodstream and heart tissue, and potentially have adverse effects on the kidneys and spleen as there were significant changes in the weights of these organs; increase in the kidneys and decrease in the spleen. Furthermore, concentrations of selenium in kidneys and iron in spleen were decreased, pointing to disruption of trace element homeostasis.ConclusionsSub-acute inhalation exposure of CuO NPs induced pulmonary inflammation, which was correlated to Cu concentrations in the lungs and started to resolve once exposure ended. Dosimetry analysis showed that Cu in the lungs was translocated into the bloodstream and heart tissue. Secondary organs affected by CuO NPs exposure were kidneys and spleen as they showed the disruption of trace element homeostasis and organ weight changes.

Safety and toxicologic evaluation of Edible Pongamia Oil: A novel food ingredient

Edible Pongamia Oil (EPO) was evaluated in an acute oral toxicity study, GLP 14-Day and 90-Day repeated dose isocaloric dietary toxicity studies in rats, and in vitro Bacterial Reverse Mutation, and in vivo Mammalian Bone Marrow Chromosome Aberration genotoxicity studies for potential use as a food ingredient. In a non-GLP acute study, an LD50 > 5000 mg/kg was determined. Subacute 14-day repeated dose dietary administration of 0, 5, 10 and 15% oil revealed no adverse changes in clinical pathology, liver histology, body weight or weight gain, food consumption or food efficiency. In a 90-day dietary study fed 0, 2.5, 5.0, 7.5 and 10.0%, no mortalities, clinical or ophthalmologic signs, body weight, body weight gain, food consumption, food efficiency or Functional Observational Battery/Motor Activity changes occurred with EPO consumption, nor were there any adverse changes in hematology, clinical chemistry, coagulation, urinalysis, or thyroid hormone values. There were no adverse macroscopic, estrus cycle, histopathologic or spermatogenesis findings, or absolute or relative organ weight changes related to administration of EPO. The No-Adverse-Effect-Level (NOAEL) was 10% in the diet, the highest dose tested, equivalent to 5163 (male) and 6469 (female) mg/kg/day in rats. No mutagenic or clastogenic genotoxic potential was reported.

Combined efficacy of Cinnamomum zeylanicum and doxorubicin against leukemia through regulation of TRAIL and NF-kappa B pathways in rat model.

Recent discoveries in cancer therapeutics have proven combination therapies more effective than individual drugs. This study describes the efficacy of the combination of Cinnamomum zeylanicum and doxorubicin against benzene-induced leukemia. Brine shrimp assay was used to assess the cytotoxicity of C. zeylanicum, doxorubicin and their combination. After AML induction in Sprague Dawley rats, the same drugs were given to rat groups. Changes in organ weight, haematological profile, and hepatic enzymes were determined. Real-time PCR was used to elucidate the effect on the expression of STMN1, GAPDH, P53 and various TRAIL and NF-kappaB components. C. zeylanicum reduced the cytotoxicity of doxorubicin. The combination treatment showed better anti-leukemic results than any of the individual drugs as evident from STMN1 expression (p < 0.001). It was particularly effective in reducing total white blood cell counts and recovering lymphocytes, monocytes and eosinophils along with hepatic enzymes ALT and AST (p < 0.001). All doses recovered relative organ weights and improved blood parameters. The combination therapy was particularly effective in inducing apoptosis, inhibition of proliferation marker GAPDH (p < 0.001) and NF-kappaB pathway components Rel-A (p < 0.001) and Rel-B (p < 0.01). Expressions of TRAIL components c-FLIP (p < 0.001), TRAIL ligand (p < 0.001) and caspase 8 (p < 0.01) were also altered. Cinnamomum zeylanicum in combination with doxorubicin helps to counter benzene-induced cellular and hepatic toxicity and improves haematological profile. The anti-leukemic effects are potentially due to inhibition of GAPDH and NF-kappa B pathway, and through regulation of TRAIL pathway. Our data suggests the use of C. zeylanicum with doxorubicin to improve anti-leukemic therapeutic regimes.

Subacute 28 days oral toxicity study of kaempferol and biochanin-A in the mouse model.

The present study was undertaken to investigate the safety of kaempferol (KEM) and biochanin-A (BCA) following subacute exposure in mice. KEM and BCA were administered in three different doses by oral administration for 28 days. Evaluation of general toxicity parameters by examining the clinical signs, body weight, organ weights, haematological, biochemical, oxidative stress parameters, and histopathology was done. Administration of KEM and BCA for 28 days did not show any clinical signs of toxicity, nor any treatment-related changes in body weight and organ weights in comparison to control. The haematological parameters such as red blood cell, white blood cell, platelets count, haemoglobin (Hb) level, haematocrit, mean corpuscular haemoglobin concentration, red cell distribution width, and platelet distribution width did not show any change in the treated groups and control. Furthermore, different biochemical parameters like markers of the liver (alanine aminotransferaseand aspartate aminotransferase), kidney (creatinine and urea), and heart (creatinine kinase-myocardial band and lactate dehydrogenase) injury along with other biochemical parameters showed nonsignificant differences between treated groups and control. Results of oxidative stress parameters in treated groups showed insignificant variations with control. The level of antioxidant enzymes such as superoxide dismutase and catalase were markedly increased in the treated groups; however, these were nonsignificant in comparison to control. In histopathology, evaluation of all vital organs, such as liver, kidney, heart, and lungs, did not show any morphological abnormalities and lesions in treated groups and control. The present study suggests that KEM and BCA have no adverse effects on the general physiology in mice.

Natural Antioxidants as a Tool to Protect the Organs from Tumors Induced Ehrlich Ascites in Mice

The objective of this study was to estimate if pre-treatment with pomegranate or beetroot juice and their combination for EAC which induces tumor in mice has the antitumor effect. Therefore, antioxidant activity DPPH scavenging was determined. The organs liver, spleen, and kidney were weighted to body weight in mice injected with EAC and treated by cisplatin, pomegranate, beetroot, and both of each pomegranate and beetroot and compared with the mice normal group. Single-cell suspension from the spleen was determined and RNA was extracted from the liver using a total RNA isolation kit and mRNA gene expression in EAC Cells was quantitative Real-Time PCR (RT-qPCR) analysis. The results showed that the pomegranate and beetroot contained high amounts of phenolic acid and flavonoid compounds and the highest antioxidant activity, which gave the highest DPPH scavenging activity. Moreover, the changes in the organ weight of vital organs were determined and compared with the control group. Because homologous recombination appears a new objective for cancer treatment by pomegranate and beetroot juice as an alternative to anticancer drugs. Therefore, pomegranate and beetroot juice have antitumor and antiproliferative influences on the Ehrlich tumor. Therefore, the pomegranate or beetroot and their combination were homologous recombinations that sensitize cells to growth inhibition and apoptosis. Keywords: Pomegranate, Beetroot juice, Ehrlich Ascites, Cisplatin, Cancer

Welfare Assessment on Healthy and Tumor-Bearing Mice after Repeated Ultrasound Imaging

Introduction: Ultrasound (US) imaging enables tissue visualization in high spatial resolution with short examination times. Thus, it is often applied in preclinical research. Diagnostic US, including contrast-enhanced US (CEUS), is considered to be well-tolerated by laboratory animals although no systematic study has been performed to confirm this claim. Therefore, the aim of this study was to screen for possible effects of US and CEUS examinations on welfare of healthy mice. Additionally, the potential influence of CEUS and molecular CEUS on well-being and therapy response to regorafenib was investigated in breast cancer-bearing mice. Material and Methods: Forty healthy Balb/c mice were randomly assigned for examination with US or CEUS (3×/week) for 4 weeks. Untreated healthy mice and mice receiving only isoflurane anesthesia served as controls (n = 10/group). Ninety-four 4T1 tumor-bearing Balb/c mice were allocated randomly to the following groups: no imaging, isoflurane anesthesia, CEUS, and molecular CEUS. They either received 10 mg/kg regorafenib or vehicle solution daily by oral gavage. Animals were examined three times within 2 weeks. CEUS measurements were performed using phospholipid microbubbles, and phospholipid microbubbles targeting the vascular endothelial growth factor receptor-2 were applied for molecular CEUS. Welfare evaluation was performed by daily observational score sheets, measuring the heart rate, Rotarod performance, and fecal corticosterone metabolites twice a week. On the last day, pathological changes in serum corticosterone concentrations, hemograms, and organ weights were obtained. Moreover, a potential influence of isoflurane anesthesia, CEUS, and molecular CEUS on regorafenib response in tumor-bearing mice was examined. Analysis of variance and Dunnett’s post hoc test were performed as statistical analyses. Results: Severity parameters were not altered after repeated US and CEUS examinations of healthy mice, but spleen sizes were significantly lower after isoflurane anesthesia. In tumor-bearing mice, no effect on animal welfare after repeated CEUS and molecular CEUS could be observed. However, leukocyte counts and spleen weights of tumor-bearing mice were significantly lower in animals examined with CEUS and molecular CEUS compared to the control groups. This effect was not visible in regorafenib-treated animals. Conclusions: Repeated US and (molecular) CEUS have no detectable impact on animal welfare in healthy and tumor-bearing mice. However, CEUS and molecular CEUS in combination with isoflurane anesthesia might attenuate immunological processes in tumor-bearing animals and may consequently affect responses to antitumor therapy.

Changes in Organ Weight, Sperm Quality and Testosterone Levels After Aluminum (Al) and Indium (In) Administration to Wistar Rats.

Aluminum and indium are widely used in industrial manufacturing, in pharmaceutical products, in medical treatments, and in food packaging, so they could reach organisms by different way. In order to clarify whether these elements are dangerous, we already demonstrated the ultrastructural modifications observed in the testicles, the epididymides, and the seminal vesicles of rat. Their pro-oxidative effect was also confirmed concomitantly to a decrease in anti-oxidant defenses in the blood, the testicles, and the liver. Thus, it seemed very logic to evaluate damages in the reproductive organs, especially on the exocrine and endocrine functions of the testicles. Aluminum and indium were intraperitoneally administered to male Wistar rats. Sperm solution was obtained from cauda epididymides. Motility, viability, density, and malformation of spermatozoa solution were assessed. Serum total unconjugated testosterone concentrations were measured using RIA technique. Our results showed a decrease in weight of the testicles, epididymides, and seminal vesicles of indium-treated rats and an increase in the weight of their kidneys. A decrease in motility, viability, and density of epididymides stored sperm as well as generation of many spermatozoa malformations was also observed especially in indium-treated rats. Testosterone levels were increased in indium but were enhanced in aluminum group. This confirmed our previous studies showing that aluminum and indium are toxic for the testicular tissues. This could be explained by the generation of reactive oxygen species (ROS) affecting strongly the exocrine and the endocrine functions of the testicles. Aluminum and indium are disturbing elements for the exocrine and endocrine functions of rat testicles.

Safety evaluation of hydroethanolic fruit extracts of Duranta erecta Linn

Background: Duranta erecta is used in traditional medicine for the treatment of myriad of diseases in most developing countries. The potential safety concerns associated with the administration of hydroethanolic fruit extracts of the plants were investigated in rats. Materials and Methods: Extracts were screened for their possible antioxidant activities by the DPPH scavenging activity. Bioactive compounds present in methanolic extracts of ripe and unripe fruits of D. erecta were identified using Gas chromatography–mass spectrometry (GC–MS). Male and female rats were grouped taking their body weights into consideration to achieve approximately equal conditions among the groups. A freshly prepared solution of DRR or DRU extract was administered orally at 100, 250 and 500 mg/kg b.wt. to different groups while normal group received distilled water daily for 28 days. Toxicity assessment was done using relative organ weight, haematological, and biochemical parameters and histological assessment. Results: The administration of extracts resulted in overall body weight increase, significant change in relative organ weight of the liver, changes in haematological index such as platelet and biochemical parameters namely ALT, AST, ALP, TBil, DBil, IBil, creatinine and urea of the tested group relative to the normal. Histological observations showed normal hepatocytes. Conclusion: The findings suggest that hydroethanolic Duranta erecta fruit extracts is safe but its prolonged use may have some level of adverse effect on the liver.

IFIT2-depleted metastatic oral squamous cell carcinoma cells induce muscle atrophy and cancer cachexia in mice.

BackgroundInterferon‐induced protein with tetratricopeptide repeat 2 (IFIT2) is a reported metastasis suppressor in oral squamous cell carcinoma (OSCC). Metastases and cachexia may coexist. The effect of cancer metastasis on cancer cachexia is largely unknown. We aimed to address this gap in knowledge by characterizing the cachectic phenotype of an IFIT2‐depleted metastatic OSCC mouse model.MethodsGenetically engineered and xenograft tumour models were used to explore the effect of IFIT2‐depleted metastatic OSCC on cancer cachexia. Muscle and organ weight changes, tumour burden, inflammatory cytokine profiles, body composition, food intake, serum albumin and C‐reactive protein (CRP) levels, and survival were assessed. The activation of the IL6/p38 pathway in atrophied muscle was measured.ResultsIFIT2‐depleted metastatic tumours caused marked body weight loss (−18.2% vs. initial body weight, P < 0.001) and a poor survival rate (P < 0.01). Skeletal muscles were markedly smaller in IFIT2‐depleted metastatic tumour‐bearing mice (quadriceps: −28.7%, gastrocnemius: −29.4%, and tibialis: −24.3%, all P < 0.001). Tumour‐derived circulating granulocyte‐macrophage colony‐stimulating factor (+772.2‐fold, P < 0.05), GROα (+1283.7‐fold, P < 0.05), IL6 (+245.8‐fold, P < 0.001), IL8 (+616.9‐fold, P < 0.001), IL18 (+24‐fold, P < 0.05), IP10 (+18.8‐fold, P < 0.001), CCL2 (+439.2‐fold, P < 0.001), CCL22 (+9.1‐fold, P < 0.01) and tumour necrosis factor α (+196.8‐fold, P < 0.05) were elevated in IFIT2‐depleted metastatic tumour‐bearing mice. Murine granulocyte colony‐stimulating factor (+61.4‐fold, P < 0.001) and IL6 (+110.9‐fold, P < 0.01) levels were significantly increased in IFIT2‐depleted metastatic tumour‐bearing mice. Serum CRP level (+82.1%, P < 0.05) was significantly increased in cachectic shIFIT2 mice. Serum albumin level (−26.7%, P < 0.01) was significantly decreased in cachectic shIFIT2 mice. An assessment of body composition revealed decreased fat (−81%, P < 0.001) and lean tissue (−21.7%, P < 0.01), which was consistent with the reduced food intake (−19.3%, P < 0.05). Muscle loss was accompanied by a smaller muscle cross‐sectional area (−23.3%, P < 0.05). Muscle atrophy of cachectic IFIT2‐depleted metastatic tumour‐bearing mice (i.v.‐shIFIT2 group) was associated with elevated IL6 (+2.7‐fold, P < 0.05), phospho‐p38 (+2.8‐fold, P < 0.05), and atrogin‐1 levels (+2.3‐fold, P < 0.05) in the skeletal muscle. Neutralization of IL6 rescued shIFIT2 conditioned medium‐induced myotube atrophy (+24.6%, P < 0.01).ConclusionsOur results suggest that the development of shIFIT2 metastatic OSCC lesions promotes IL6 production and is accompanied by the loss of fat and lean tissue, anorexia, and muscle atrophy. This model is appropriate for the study of OSCC cachexia, especially in linking metastasis with cachexia.

Phytochemical identification, acute and subchronic oral toxicity assessments of hydroalcoholic extract of Acroptilon repens in BALB/c mice: A toxicological and mechanistic study

Acroptilon repens (L.) DC, commonly known as Rhaponticum repens, is a popular traditional phytomedicine. The current study was conducted to evaluate the acute and subchronic toxicity of the hydroalcoholic extract of this herb with regard to its terpenoid contents in a BALB/c mice model and to investigate the toxicity of this medicinal herb. Identification of extract components of the plant was done using gas chromatography (GC)-mass spectrometry. In order to establish the acute toxicity model, a single dose of 2000 mg/kg of the extract was given orally to male mice and in the subchronic toxicity study, the extract was consecutively administered at doses 250, 500, and 1000 mg/kg for 28 days. After 28 and 42 days, signs of toxicity and mortality were observed. Organ weight changes and the toxicity-associated parameters such as biochemical indicators, oxidative stress indices, mitochondrial parameters, apoptosis-associated gene expression levels, and pro-inflammatory cytokines were evaluated along with the histopathological examination. GC analysis showed that the terpenoids are the major components of the extract. The LD50 value (2 g/kg) was obtained in the acute toxicity assay; the subchronic administration caused a significant elevation in the serum biomarkers as well as in the levels of lipid peroxidation, protein carbonyl, and ROS. Besides, significant reductions in the superoxide dismutase and catalase activities were observed. This toxic effect was further confirmed by histological studies, cytokine assay, and gene expression assays. Following the treatment discontinuation, the abnormalities in the values of biochemical parameters and histopathological changes returned to normal. These findings demonstrate that the subchronic administration of the hydroalcoholic extract of A. repens can reversibly cause toxicity by inducing oxidative stress and mitochondrial dysfunction.