Polysaccharide Fraction from Campomanesia adamantium and Campomanesia pubescens Attenuates 5-Fluorouracil-Induced Intestinal Mucosal Inflammation in Mice.

Gastrointestinal toxicity, including diarrhea and inflammation, is commonly observed with the use of 5-fluorouracil (5-FU). Several studies have shown that polysaccharides are interesting bioactive macromolecules for the treatment or prevention of gastrointestinal diseases. Therefore, in this study, the effect of a polysaccharide fraction from a mixture of two Guavira species (Campomanesia adamantium and Campomanesia pubescens), referred to here as CPW, on the development of intestinal mucositis was investigated. Intestinal mucositis was induced by a single injection of 5-FU (450 mg/kg), and various doses of CPW (3-100 mg/kg) were tested. CPW attenuated disease development and prevented small bowel dysmotility and colon shortening. CPW prevented the increase in villi width, crypt depth, and mucosal thickness in the duodenum, but not in the colon. Preservation of mucus, reduction of oxidative stress, inflammation, and prevention of the 5-FU-induced enlargement and swelling of the spleen were observed. In conclusion, this study demonstrated for the first time that CPW alleviates the intestinal damage induced by 5-FU and could be used as an adjuvant strategy during chemotherapy.

Development of a self-limiting model of methotrexate-induced mucositis reinforces butyrate as a potential therapy.

Gastrointestinal mucositis is a complication of anticancer treatment, with few validated in vitro systems suitable to study the complex mechanisms of mucosal injury. Therefore, we aimed to develop and characterize a chemotherapeutic-induced model of mucositis using 3D intestinal organoids. Organoids derived from mouse ileum were grown for 7 days and incubated with different concentrations of the chemotherapeutic agent methotrexate (MTX). Metabolic activity, citrulline levels and cytokine/chemokine production were measured to determine the optimal dosage and incubation time. The protective effects of folinic acid on the toxicity of MTX were investigated by pre-treating organoids with (0.0005-50 µg/mL) folinic acid. The impact of microbial-derived short-chain fatty acids was evaluated by supplementation with butyrate in the organoid model. MTX caused a dose-dependent reduction in cell metabolic activity and citrulline production that was salvaged by folinic acid treatment. Overall, MTX causes significant organoid damage, which can be reversed upon removal of MTX. The protective effect of folinic acid suggest that the organoids respond in a clinical relevant manner. By using the model for intervention, it was found that prophylactic treatment with butyrate might be a valuable strategy for prophylactic mucositis prevention.

Cytoprotective effects of spleen-invigorating pill against 5-fluorouracil injury to mouse bone marrow stromal cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Spleen-invigorating pills (SIP) are composed of Codonopsis, fried Atractylodes, tangerine peel, Fructus aurantii immaturus (fried), fried hawthorn, and colored malt. SIP strengthens the spleen and increases appetite and is often used as a chemotherapy adjuvant. AIM OF THE STUDY: We aimed to explore the protective effects and mechanism of action for SIP on mouse bone marrow stromal cells (OP9) injured by 5-fluorouracil (5-FU). MATERIALS AND METHODS: The effects of SIP on OP9 cells injured by 5-FU were evaluated, and high-performance liquid chromatography (HPLC) was used as a quality control method. The experiments were divided into a control group, a model group, an epidermal growth factor (EGF) treatment group, and an SIP treatment group. The cell survival rate, apoptotic cell morphology, cell apoptosis rate, and the contents of caspase 3 were evaluated to determine the protective effects of SIP in OP9 cells injured by 5-FU. Network pharmacology was used to predict the mechanism through which SIP mediates anti-chemotherapy damage. The nitric oxide (NO) and nitric oxide synthase (iNOS) levels and the expression of nuclear factor erythroid-2 related factor 2 (Nrf2) and p62 protein were detected to explore the mechanism through which SIP mediates anti-chemotherapy damage through the regulation of oxidative stress. RESULTS: Cell counting kit-8 (CCK8) detection showed that 5-FU reduced OP9 cell survival, and SIP blocked the inhibition of OP9 cell growth induced by 5-FU. When OP9 cells were treated with both SIP (10 g L-1) and 5-FU (2.5 × 10-2 g L-1) for 24 h, compared with the model group, the early apoptosis rates significantly decreased, and the activity of caspase 3 was significantly reduced. The results of network pharmacology and Western blot showed that compared with the model group, in the SIP group, the NO levels decreased, iNOS release decreased, and the expression of Nrf2 and p62 proteins increased. CONCLUSION: The protective effects of SIP on OP9 cells injured by 5-FU were significant. SIP may play a cytoprotective role by mediating changes in oxidative stress-related proteins. The specific mechanism of action through which SIP mediates these effects remains to be further studied.

Metformin and omega-3 fish oil elicit anti-inflammatory effects via modulation of some dysregulated micro RNAs expression and signaling pathways in experimental induced arthritis.

OBJECTIVE: Rheumatoid arthritis is a progressive inflammatory disease with multiple dysfunctional intracellular signaling pathways that necessitate new approaches for its management. Hence, the study aimed to inspect the ability of the combination therapy of metformin and omega-3 to modulate different signaling pathways and micro RNAs such as (miR-155, miR-146a and miR-34) as new targets in order to mitigate adjuvant-induced arthritis and compare their effect to that of methotrexate. METHODS: Fourteen days post adjuvant injection, Sprague-Dawley rats were treated orally with metformin (200 mg/kg/day) and/or omega-3 (300 mg/kg/day) or intraperitoneally with methotrexate (2 mg/kg/week) for 4 weeks. RESULTS AND CONCLUSION: All drug treatments amended the arthrogram score and hind paw swelling as well as decreased serum tumor necrosis factor (TNF)-α and interleukin (IL)-1ß levels. On the molecular level, all therapies activated phospho-5'adenosine monophosphate-activated protein kinase (p-AMPK) and protein phosphatase 2A (PP2A), while they inhibited phospho-mammalian target of rapamycin (p-mTOR), phospho-signal transducers and activators of transcription (p-STAT3), nuclear factor (NF)-κB p65 subunit, phosho38 mitogen-activated protein kinase (p38 MAPK) and phospho- c-Jun N-terminal kinase (p-JNK). In addition, they decreased the elevated expression level of miRNA-155, 146a and increased the expression level of miRNA-34 and they decreased the expression level of retinoic acid receptor related orphan receptor γT (RORγT) and increased that of fork head box P3 (FOXP3), correcting Th17/Treg cells balance. On most of the aforementioned parameters, the effect of the combination therapy was comparable to that of methotrexate, emphasizing that this combination possesses better additive anti-inflammatory effect than either drug when used alone. In addition, the combination was capable of normalizing the serum transaminases levels as compared to untreated group offering hepatoprotective effect and suggesting the possibility of its use as a replacement therapeutic strategy for MTX in rheumatoid arthritis.

Effects of vitamin B12 on methotrexate hepatotoxicity: evaluation of receptor-interacting protein (RIP) kinase.

In the study, we aimed to show the effects of vitamin B12 on the necrosis caused by methotrexate (MTX), a folic acid antagonist. Thirty-two rats were randomly assigned to four groups of eight rats per group. Control (n = 8), Vit B12 (n = 8) 3 µg/kg/ip B12 (15 days) per day throughout the experiment, MTX (n = 8) injected with a single dose of 20 mg/kg/ip MTX on 8th day of experiment, MTX + Vit B12 (n = 8) injected with a single dose of 20 mg/kg ip methotrexate on 8th day of experiment + 3 µg/kg/ip Vit B12 (15 days) per day throughout the experiment. Oxidant (TOS)/antioxidant (TAS) system, TNF-α and TGF-ß levels, AST and ALT, serum vitamin B12 levels were determined in the tissue. Cyclooxygenase-2 (Cox-2), receptor-interacting protein kinase 1 (RIP1) and 3 (RIP3) immunohistochemistry were applied to the liver tissue. TOS increased; TAS decreased; TNF-α and TGF-ß levels increased; AST and ALT levels changed after MTX hepatotoxicity. Vit B12 decreased significantly. COX-2, RIP1, and RIP3 immunoreactivity increased. Vit B12 showed improvement in all of the negative results. Vit B12 is an important supplement to be used against necrosis in tissue after MTX hepatotoxicity.

Synthesis and characterization of thermo/pH-sensitive pectin-graft-poly(dimethylaminoethyl methacrylate) coated magnetic nanoparticles.

Herein, thermo- and pH-sensitive pectin-graft-poly(dimethylaminoethyl methacrylate) copolymer-coated magnetic nanoparticles were synthesized via a green and rapid synthetic approach based on microwave irradiation. Firstly, a novel thermo- and pH-sensitive pectin-graft-poly(dimethylaminoethyl methacrylate) copolymer (Pec-g-PolyDMAEMA) was synthesized and then, Pec-g-PolyDMAEMA based magnetic nanoparticles (Pec-g-PolyDMAEMA@Fe3O4) were produced via microwave-assisted co-precipitation method. The thermo/pH/magnetic field multi-sensitive hybrid nanoparticle was characterized by techniques like TEM, VSM, FT-IR, and TGA/DSC. In vitro release studies of 5-Fluorouracil (FL) were carried out by altering the temperature (37 and 44°C), pH (5.5 and 7.4) and presence of an AMF. The FL release of Pec-g-PolyDMAEMA@Fe3O4@FL exhibited pH-sensitive behavior. They showed thermo/pH-sensitive FL release features with the greatest release of FL at 37°C (56%) than at 44°C (40%) and at pH of 7.4 (63%) than at pH of 5.5 (45%) within 48h. The FL release was also significantly increased (100%) with the presence of a 50 mT magnetic field. These results indicate that the developed Pec-g-PolyDMAEMA@Fe3O4 nanoparticles are promising in the application of multi-stimuli-sensitive delivery of drugs.

Protective effects of Glycyrrhiza glabra supplementation against methotrexate-induced hepato-renal damage in rats: An experimental approach.

ETHNOPHARMACOLOGICAL RELEVANCE: In traditional medicine, Glycyrrhiza glabra, commonly known as liquorice, is known to possess promising pharmacological properties including anti-oxidative, anti-inflammatory, gastro, hepato and nephro-protective activities. AIM: The present study investigated the protective effects of Glycyrrhiza glabra rhizome extract (GGE) on MTX-induced hepato-renal damage in Wistar albino rats. MATERIALS AND METHODS: Rats were pre-treated with GGE (100, 200 or 400 mg/kg) from day 1 to 15 and administered MTX (20 mg/kg) on day 4. Methotrexate-induced hepato-renal damage was assessed by serum toxicity biomarkers (AST, ALT, BUN and creatinine), oxidative stress estimation (MDA, GSH, SOD, CAT, peroxidase and glutathione reductase), interleukins profiling (TNF-α, IL-1ß, IL-6 and IL-12), tissue histopathology and immunohistochemical (caspase-3 and NFkB) examination. RESULTS: MTX induced hepato-renal damage resulted in elevated serum levels of AST, ALT, BUN and creatinine, increased pro-inflammatory cytokines concentration and accumulation of MDA and reduced levels of GSH, SOD, CAT, peroxidase and glutathione reductase. Conversely, co-treatment with GGE dose-dependently ameliorated oxidative stress, serum interleukins, hepato-renal toxicity biomarkers (p < 0.001), preserved tissue architecture and downregulated both caspase-3 and NFkB expression in hepato-renal tissue. CONCLUSION: The above results suggested that GGE can alleviate MTX-induced hepato-renal damage by decreasing oxidative stress and suppressing the ensuing activation of pro-apoptotic and pro-inflammatory pathways.

Role of nitric oxide donor in methotrexate-induced testicular injury via modulation of pro-inflammatory mediators, eNOS and P-glycoprotein.

Methotrexate (MTX) is a widely used chemotherapeutic agent but its clinical use is challenged with different forms of toxicities including testicular injury. The aim of the current study was to evaluate the potential protective effect of potassium channel opener, nicorandil (NIC) (3 and 10 mg/kg/day) on MTX-induced testicular injury in a rat model. Rats were randomly divided into four groups (nine rats each) and treated for 2 weeks as follows: (I) normal control (CON group) received vehicle, (II) model group (MTX group) given MTX (20 mg/kg) single intraperitoneal (i.p.) injection dose on 11th day, (III) MTX + NLD group treated with NIC (3 mg/kg/day) orally for 2 weeks and MTX (20 mg/kg) single i.p. dose on 11th day, and (IV) MTX + NHD group treated with NIC (10 mg/kg/day) orally for 2 weeks and MTX (20 mg/kg) single i.p. injection on the 11th day. The testicular injury was assessed biochemically via serum testosterone, total antioxidant capacity, testicular oxidative stress parameters, P-glycoprotein, tumor necrosis factor-alpha, and interleukin-1ß. Furthermore, histopathological evaluation, endothelial nitric oxide synthase (eNOS) immunoexpression, and detection of p53 expression level using Western blotting were performed. Results showed that MTX induced testicular injury which was proved by both biochemical and histopathological evaluations. Our results concluded that NIC pretreatment attenuated MTX-induced testicular injury via significantly increased eNOS immunoexpression, antiapoptotic, anti-inflammatory, and antioxidant properties. Interestingly, NIC high dose is more protective than low dose.

Ready to go 3D? A semi-automated protocol for microwell spheroid arrays to increase scalability and throughput of 3D cell culture testing.

3-dimensional (3D) cell cultures are being increasingly recognized as physiologically more relevant in vitro models than traditional monolayer cultures, because they better mimic in vivo-like microenvironment, cell-cell and cell-extracellular matrix interactions. Nevertheless, the broader use of 3D models might be limited by requirements for special consumables, equipment, or skills for 3D cell cultures, and by their limited throughput and scalability. In this study, we optimized and adapted a commercially available agarose-micromolding technique to produce scaffold-free spheroid cultures. Brightfield microscopy was used for routine nondestructive and noninvasive evaluation of spheroid formation and growth. The workflow is compatible with manual, as well as high speed automated microscopic image acquisition, and it is supplemented with an in-house developed macro 'Spheroid_Finder' for open source software Fiji to facilitate rapid automated image analysis. This protocol was used to characterize and quantify spheroid formation and growth of two different hepatic cell lines, hTERT immortalized, but non-cancerous, adult human liver stem cell line HL1-hT1, and human hepatocellular carcinoma cell line HepG2, as well as their responses to a model antiproliferative and cytotoxic agent, 5-fluorouracil. The complete protocol provides a simple and ready-to-use solution to initiate scaffold-free spheroid cultures in any laboratory with standard equipment for mammalian in vitro cell culture work. Thus, it allows to increase throughput and scale of spheroid culture experiments, which can be greatly utilized in different areas of biomedical, pharmaceutical and toxicological research.

Patchouli alcohol attenuates 5-fluorouracil-induced intestinal mucositis via TLR2/MyD88/NF-kB pathway and regulation of microbiota.

Intestinal mucositis causes great suffering to cancer patients who undergo chemotherapy and radiotherapy. Owing to the uncertain side effects of anticancer drugs to attenuate patients' intestinal mucositis, many studies focused on traditional Chinese medicine (TCM). Patchouli alcohol (PA) is an active compound extracted from Pogostemon cablin, and has potent gastrointestinal protective effect. However, whether PA has an effect on intestinal mucositis is still unknown. Therefore, we established a rat model of intestinal mucositis via intraperitoneal injection of 5-fluorouracil, and intragastrically administrated PA (10, 20, and 40 mg/kg) to evaluate the effect of PA on intestinal mucositis. The routine observation (body weight, food intake, and diarrhea) in rats was used to detect whether PA had an effect on intestinal mucositis. Levels of inflammatory cytokines (TNF-α, IL-1ß, IL-6, IL-10, and MPO), mucosal barrier proteins (zonula occludens -1 (ZO-1), claudin-1, occludin, myosin light chain (MLC), and mucin-2) and intestinal microbiota were determined to elucidate the underlying mechanism of PA action on intestinal mucositis in rats. The results showed that PA could effectively improve body weight, food intake, and diarrhea in intestinal mucositis rats, preliminary confirming PA efficacy. Further experiments revealed that PA not only decreased the levels of TNF-α, IL-1ß, IL-6, and MPO but also increased the level of IL-10 significantly. In addition, the expression of mucosal barrier proteins and microbiota community were also improved after PA treatment in diseased rats. Hence, PA may prevent the development and progression of intestinal mucositis by improving inflammation, protecting mucosal barrier, and regulating intestinal microbiota.

Patchouli oil ameliorates 5-fluorouracil-induced intestinal mucositis in rats via protecting intestinal barrier and regulating water transport.

ETHNOPHARMACOLOGICAL RELEVANCE: Pogostemon cablin, commonly named "Guang-Huo-Xiang" in China, has long been renowned for its ability to dispel dampness and regulate gastrointestinal functions. Patchouli oil (P.oil), the major active fraction of Pogostemon cablin, has been traditionally used as the principal component of Chinese medicinal formulae to treat exterior syndrome and diarrhea. However, the effects of P.oil in treating 5-fluorouracil (5-FU)-induced intestinal mucositis have not yet been reported. AIM OF THE STUDY: To investigate the protective effects of P.oil against 5-FU-induced intestinal mucositis and the mechanisms underlying these effects. MATERIALS AND METHODS: Sprague-Dawley rats were intraperitoneally injected with 5-FU (30 mg/kg) to establish an intestinal mucositis model. Meanwhile, rats with intestinal mucositis were orally administered with P.oil (25, 50, and 100 mg/kg). Histological analysis, ELISA (for detecting inflammatory cytokines and aquaporins), immunohistochemistry analysis (for examining caspases), qRT-PCR analysis (for assessment tight junctions), and western blotting analysis (for the assessment of TLR2/TLR4-MyD88 and VIP-cAMP-PKA signaling pathway-related proteins) were performed to estimate the protective effects of P.oil against intestinal mucositis and the mechanisms underlying these effects. RESULTS: The histopathological assessment preliminarily exhibited that P.oil alleviated the 5-FU-induced damage to the intestinal structure. After P.oil administration, the elevation of the expression of cytokines (TNF-α, IFN-γ, and IL-13) decreased markedly and the activation of NF-κB and MAPK signaling was significantly inhibited. P.oil also increased the mRNA expression of ZO-1 and Occludin, thereby stabilizing intestinal barrier. In addition, P.oil decreased the expressions of caspase-8, caspase-3, and Bax, and increased the expression of Bcl-2, thereby reducing the apoptosis of the intestinal mucosa. These results were closely related to the regulation of the TLR2/TLR4-MyD88 signaling pathway. It has been indicated that P.oil possibly protected the intestinal barrier by reducing inflammation and apoptosis. Furthermore, this study showed that P.oil inhibited the abnormal expression of AQP3, AQP7, and AQP11 by regulating the VIP-cAMP-PKA signaling pathway. Furthermore, it restored the intestinal water absorption, thereby alleviating diarrhea. CONCLUSIONS: P.oil ameliorated 5-FU-induced intestinal mucositis in rats via protecting intestinal barrier and regulating water transport.

Thyme (Thymus vulgaris) leaf extract modulates purinergic and cholinergic enzyme activities in the brain homogenate of 5-fluorouracil administered rats.

This study investigated the effect of Thyme (Thymus vulgaris) leaf aqueous extract (TE) on purine and acetylcholine hydrolyzing enzyme activities and antioxidant status in the brain of 5-fluorouracil (5-FLU) administered rats. Sixty adult Wistar rats (210-225 g) were divided into Ten groups (n = 6). Group 1: received normal saline orally (NC), group 2, 3, 4 and 5 rats were respectively received 100, 200 and 400 mg/kg of TE and Vitamin C (25 mg/kg) orally; group 6 rats received normal saline orally and 150 mg/kg of 5-FLU via i.p., on the last day; rats in group 7, 8, 9 and 10 were orally pretreated with 100, 200 and 400 mg/kg of TE and Vit C (25 mg/kg) respectively, and administered with 150 mg/kg of 5-FLU i.p., on the last day. The purine [adenosine 5'-tri- and di-phosphatases (ATPase and ADPase), adenosine deaminase (ADA)] nucleotide and acetylcholine hydrolyzing [acetylcholinesterase (AChE) and butrylcholinesterase (BChE)] enzymes as well as arginase activities and antioxidant status were assessed. The result revealed that untreated 5-FLU rats had increased enzyme activities, which was reduced in the normal rats treated TE compared to NC. Interestingly, the TE-pretreated rats administered with 5-FLU had reduced enzyme activities compared with untreated 5-FLU rats. The antioxidant status was also normalized in the 5-FLU administered rats pretreated with TE Thus, the neuroprotective mechanisms of TE could be linked to attenuation of neuroactive enzymes and antioxidant status.

Agarwood Extract Mitigates Intestinal Injury in Fluorouracil-Induced Mice.

Agarwood is used to treat gastrointestinal diseases. Although our previous studies demonstrated that agarwood ethanol extract produced by the whole-tree agarwood-inducing technique (WTAAE) improves intestinal peristalsis, the intestinal protective effect of WTAAE remains unclear. This study aimed to evaluate the protective effect of WTAAE on the intestinal injury induced by fluorouracil (5-FU) and explore its potential mechanism. Institute of Cancer Research (ICR) mice were given agarwood ethanol extracts (AAEs) (details in materials part), including WTAAE (0.71, 1.42 and 2.84 g/kg), wild agarwood ethanol extract (WAAE) and burning-chisel-drilling agarwood ethanol extract (FBAAE) (2.84 g/kg). A colon injury model was induced by 5-FU. After 14 d of treatment, the histopathology and biochemical and molecular parameters were measured. Our results indicated that WTAAE enhanced the intestinal advancing rate and alleviated the severity of colon injury similar the WAAE and better than FBAAE. Simultaneously, WTAAE reduced the nitric oxide (NO) concentration and increased the glutathione (GSH) and superoxide dismutase (SOD) levels. WTAAE also reduced the levels of interleukin-17 (IL-17) and IL-33 and elevated the level of IL-10. Furthermore, WTAAE upregulated the mRNA expression of the nuclear factor-E2-related factor 2-antioxidant response element (Nrf2-ARE) pathway and downregulated the mRNA levels of the nuclear factor-kappaB (NF-κB) pathway. WTAAE had a mitigating effect on intestinal damage, suggesting that it could be used as an intestinal protective and adjuvant therapy drug for intestinal injury induced by chemical drugs.

Treatment with selenium-enriched Saccharomyces cerevisiae UFMG A-905 partially ameliorates mucositis induced by 5-fluorouracil in mice.

PURPOSE: Gastrointestinal mucositis is a major problem associated with cancer therapy. To minimize these deleterious effects, simultaneous administration of antioxidant components, such as selenium, can be considered. There is a growing interest in the use of yeasts because they are able to convert inorganic selenium into selenomethionine. In the present study, oral administration of Saccharomyces cerevisiae UFMG A-905 enriched with selenium was evaluated as an alternative in minimizing the side effects of 5FU-induced mucositis in mice. METHODS: Mice body weight, food consumption, faeces consistency and the presence of blood in faeces were assessed daily during experimental mucositis induced by 5-fluorouracil (5FU). Blood was used for intestinal permeability determination, and small intestine for oxidative stress, immunological and histopathological examination. RESULTS: The increased intestinal permeability observed with mucositis induction was partially reverted by S. cerevisiae and selenium-enriched yeast. Both treatments were able to reduce myeloperoxidase activity, but only selenium-enriched yeast reduced eosinophil peroxidase activity. CXCL1/KC levels, histopathological tissue damage and oxidative stress (lipid peroxidation and nitrite production) in the small intestine were reduced by both treatments; however, this reduction was always higher when treatment with selenium-enriched yeast was evaluated. CONCLUSIONS: Results of the present study showed that the oral administration of S. cerevisiae UFMG A-905 protected mice against mucositis induced by 5-FU, and that this effect was potentiated when the yeast was enriched with selenium.

Gemcitabine loaded microbubbles for targeted chemo-sonodynamic therapy of pancreatic cancer.

Pancreatic cancer remains one of the most lethal forms of cancer with a 10-year survival of <1%. With little improvement in survival rates observed in the past 40 years, there is a significant need for new treatments or more effective strategies to deliver existing treatments. The antimetabolite gemcitabine (Gem) is the most widely used form of chemotherapy for pancreatic cancer treatment, but is known to produce significant side effects when administered systemically. We have previously demonstrated the benefit of combined chemo-sonodynamic therapy (SDT), delivered using oxygen carrying microbubbles (O2MB), as a targeted treatment for pancreatic cancer in a murine model of the disease. In this manuscript, we report the preparation of a biotin functionalised Gem ligand for attachment to O2MBs (O2MB-Gem). We demonstrate the effectiveness of chemo-sonodynamic therapy following ultrasound-targeted-microbubble-destruction (UTMD) of the O2MB-Gem and a Rose Bengal loaded O2MB (O2MB-RB) as a targeted treatment for pancreatic cancer. Specifically, UTMD using the O2MB-Gem and O2MB-RB conjugates reduced the viability of MIA PaCa-2, PANC-1, BxPC3 and T110299 pancreatic cancer cells by >60% (p < 0.001) and provided significant tumour growth delay (>80%, p < 0.001) compared to untreated animals when human xenograft MIA PaCa-2 tumours were treated in SCID mice. The toxicity of the O2MB-Gem conjugate was also determined in healthy non-tumour bearing MF1 mice and revealed no evidence of renal or hepatic damage. Therefore, the results presented in this manuscript suggest that chemo-sonodynamic therapy using the O2MB-Gem and O2MB-RB conjugates, is potentially an effective targeted and safe treatment modality for pancreatic cancer.

Preclinical evaluation of NUDT15-guided thiopurine therapy and its effects on toxicity and antileukemic efficacy.

Thiopurines (eg, 6-mercaptopurine [MP]) are highly efficacious antileukemic agents, but they are also associated with dose-limiting toxicities. Recent studies by us and others have identified inherited NUDT15 deficiency as a novel genetic cause of thiopurine toxicity, and there is a strong rationale for NUDT15-guided dose individualization to preemptively mitigate adverse effects of these drugs. Using CRISPR-Cas9 genome editing, we established a Nudt15-/- mouse model to evaluate the effectiveness of this strategy in vivo. Across MP dosages, Nudt15-/- mice experienced severe leukopenia, rapid weight loss, earlier death resulting from toxicity, and more bone marrow hypocellularity compared with wild-type mice. Nudt15-/- mice also showed excessive accumulation of a thiopurine active metabolite (ie, DNA-incorporated thioguanine nucleotides [DNA-TG]) in an MP dose-dependent fashion, as a plausible cause of increased toxicity. MP dose reduction effectively normalized systemic exposure to DNA-TG in Nudt15-/- mice and largely eliminated Nudt15 deficiency-mediated toxicity. In 95 children with acute lymphoblastic leukemia, MP dose adjustment also directly led to alteration in DNA-TG levels, the effects of which were proportional to the degree of NUDT15 deficiency. Using leukemia-bearing mice with concordant Nudt15 genotype in leukemia and host, we also confirmed that therapeutic efficacy was preserved in Nudt15-/- mice receiving a reduced MP dose compared with Nudt15+/+ counterparts exposed to a standard dose. In conclusion, we demonstrated that NUDT15 genotype-guided MP dose individualization can preemptively mitigate toxicity without compromising therapeutic efficacy.

Changes in cognition and dendritic complexity following intrathecal methotrexate and cytarabine treatment in a juvenile murine model.

Acute lymphoblastic leukemia (ALL) is the most prevalent childhood cancer and accounts for 26.8% of cancer diagnoses among children, worldwide-approximately 3000 children each year. While advancements in treating ALL have led to a remission rate of more than 90%, many survivors experience adverse neurocognitive and/or neurobehavioral effects as a result of intrathecal chemotherapy. Methotrexate (MTX) is commonly administered with cytosine arabinoside (AraC, cytarabine) during intrathecal chemotherapy for ALL. To date, few studies exist that test the cognitive effects of intrathecal injections of MTX/AraC on juvenile populations. The purpose of our study was to investigate the combined effects of MTX/AraC on cognition and dendritic structure in the hippocampus in juvenile male mice. Twenty, 21-day-old male C57BL/6 mice were used in this study; 10 mice received intrathecal MTX/AraC treatment, and 10 were given intrathecal saline injections. Five weeks after injections, we tested the animals' hippocampus-dependent cognitive performance in the Morris water maze. After the first day of hidden-platform training, we observed that the mice that received MTX/AraC treatment showed signs of significant impairment in spatial memory retention. MTX/AraC treatment significantly compromised the dendritic architecture and reduced mushroom spine density in the dorsal ganglion (DG), CA1, and CA3 areas of the hippocampus. The present data provided evidence that MTX/AraC compromised the dendritic architecture and impaired hippocampal dependent cognition. This could provide insight into chemotherapy-induced cognitive decline in juvenile patients treated for ALL.

Hepatoprotective effect of berberine against methotrexate induced liver toxicity in rats.

Hepatotoxicity is one of the major side effects of methotrexate (MTX), which restricts the clinical use of this drug. Berberine (BBR) is a natural compound with multiple pharmacological activities such as antioxidant, antiapoptotic and anti-inflammatory effects. In this study, the effect of BBR on MTX-induced hepatotoxicity was studied. A total number of 28 male Wistar rats were randomly divided into four experimental groups. Rats were pretreated with BBR orally with dose of 100mg/kg for 10 consecutive days and MTX (20mg/kg, intraperitoneally) was administrated on the 9th day. Then on day 11, blood samples were collected to determine serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP). The extracted livers were used for histological examination, biochemical assays and real time PCR studies. Malondialdehyde (MDA), glutathione (GSH), protein carbonyl (PC), nitric oxide (NO) levels, catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx) and myeloperoxidase (MPO) activities were assessed in hepatic tissue. In addition, the expression of SOD and PGx was measured using real-time PCR method in hepatic tissue. Results showed that MTX administration significantly increases AST, ALT and ALP levels (all p<0.001). It also, increases MDA, PC, NO levels and MPO activity (p<0.001, p<0.01, p<0.05 and p<0.01 respectively). Moreover, MTX decreases hepatic GSH level, SOD, GPx and CAT activities (all p<0.001). Pre-treatment with BBR for 10days prevented some of these changes. Serum levels of AST and ALT decreased (all p<0.001). Hepatic MDA level decreased (p<0.001) and GSH level as well as GPx activity increased (p<0.05 and p<0.01 respectively). Our results indicated that BBR might be useful for prevention of the hepatotoxicity induced by MTX via ameliorative effects on biochemical and oxidative stress indices.

Plant Polysaccharides Attenuate Fluorouracil Toxicity for the Small Intestinal Epithelium.

Polysaccharides from Tussilago farfara L., Acorus calamus L., and Echinacea purpurea (L.) Moench attenuated the toxic effect of fl uorouracil on the small intestinal epithelium of mice with Lewis lung carcinoma. Addition of polysaccharides to chemotherapy protocols stimulated reparative regeneration processes in the small intestine damaged by the cytostatic treatment. No stimulating effects of the polysaccharides on tumor growth and metastasizing were revealed.

Protective effect of Bu-Zhong-Yi-Qi decoction, the water extract of Chinese traditional herbal medicine, on 5-fluorouracil-induced intestinal mucositis in mice.

Intestinal mucositis is a serious toxic side effect of 5-fluorouracil (5-FU) treatment. Bu-Zhong-Yi-Qi decoction (BZYQD), a water extract of Chinese traditional herbal medicine, is widely used in chemotherapy in Asia as an alternative treatment to reduce the side effects of chemotherapy. However, the mechanism is unknown. To evaluate its mechanism, we investigated the effect of BZYQD on 5-FU-induced intestinal mucositis in mice, especially with regard to apoptosis in the intestinal mucosal epithelia. In the present study, mice were divided into three groups: control, 5-FU, and 5-FU + BZYQD. Mice in the 5-FU and 5-FU + BZYQD groups were administered 5-FU (100 mg/kg/day, intraperitoneally) for 6 days, and the mice in the latter group were given BZYQD (8 g/kg/day, intragastrically) beginning 4 days before 5-FU and continuing until the termination of the experiment. Loss in body weight and diarrhea during the 5-FU treatment were significantly attenuated by administration of BZYQD. The morphological signs of intestinal damage, including shortened villi height, crypt destruction, apoptosis, and necrosis, in intestinal mucosal epithelia were also reversed, accompanied by reduced neutrophil infiltration, nitrite levels, and inflammatory factors (tumor necrosis factor α and interleukin 1ß) and increased levels of reduced glutathione. These results suggest that BZYQD inhibits 5-FU-induced intestinal mucositis, and this effect may be due to the reduction in apoptosis and necrosis in intestinal mucosal epithelia via the suppression of inflammatory cytokine upregulation. In conclusion, inhibiting cytokine-mediated apoptosis or necrosis can be the molecular mechanism by which BZYQD reduces the gastrointestinal side effects of cancer chemotherapy.