Methotrexate In Rats Research Articles

The Possible Ameliorative effect of Zafirlukast on Renal Toxicity Induced by Methotrexate in Experimental Animals

Methotrexate is a chemotherapeutic agent. Nephrotoxicity is an important side effect of treatment with methotrexate. Zafirlukast is a cystenyl leukotriene antagonist that acts as an anti-inflammatory and antioxidant agent. The aim of this study was to analyze the nephrotoxicity induced by methotrexate in rats and to evaluate the possible ameliorative effect of zafirlukast. Ethanol (1ml/kg, orally once daily) and zafirlukast (40mg/kg, orally once daily) were given for 10 days. Methotrexate (20mg/kg I.P, single injection) and, zafirlukast (40mg/kg, orally once daily) was given 5 days before methtrexate and then 5 days after the chemotherapeutic agent. Rats will be sacrificed after the 10th day from methotrexate intake. Rats given methotrexate alone had significantly higher MDA (malondialdehyde), lower reduced glutathione, catalase, superoxide dismutase in kidney tissue and higher blood urea nitrogen and serum creatinine levels than the control groups. Co-treatment of zafirlukast with methotrexate had an ameliorative effect on the previous parameters. Light & electronic microscopic pictures revealed that methotrexate caused marked degenerative changes in the kidney of rats. Restoration of the normal architecture of kidney tissues were observed in rats received methotrexate with zafirlukast. Thus, methotrexate can induce oxidative damage in rat kidney & zafirlukast has an ameliorative effect against this damage

The effects of lycopene on intestinal injury due to methotrexate in rats

Objective: The aim of this study was to investigate the effects of lycopene (Lyc) on methotrexate (Mtx)-induced intestinal damage in rats.Method: Twenty-eight male Sprague Dawley rats were divided into four equal groups: control, Mtx, Lyc, and Mtx-L.Control group: Rats were given only the vehicle.Lyc group: Rats were given Lyc (10 mg/kg) with corn oil by oral gavage for 10 days.Mtx group: Rats were injected intraperitoneally with a single dose of 20 mg/kg of Mtx and given corn oil by oral gavage.Mtx-L group: Rats were treated with Lyc (10 mg/kg) for 10 days after a single dose of Mtx (20 mg/kg). All of the rats were euthanized using terminal anesthesia, and the intestinal tissues were removed for histological examination and for pro-inflammatory cytokine measurement (tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β)), total oxidative status (TOS), total antioxidant capacity (TAC), and oxidative stress index (OSI).Results: Mtx administration increased histopathological damage and increased TNF-α, IL-1β, TOS, TAC, and OSI levels in the small intestine tissues. Lyc therapy applied to the Mtx-L group provided significant improvement in all parameters of histopathological damage to the small intestine and significantly reduced the levels of IL-1β, TOS, and OSI in the intestinal tissues.Conclusions: The results of this study indicate that Lyc might be useful for protecting intestinal damage induced by Mtx in rats by reducing the increased oxidative stress and pro-inflammatory cytokine (IL-1β) levels.

Effects of nimesulide on the small intestine mucositis induced by methotrexate in rats.

Intestinal mucositis is one of the major problems in the patients receiving cancer treatment. Nimesulide is a drug with antioxidant, antiinflammatory and antiulcer features. We aimed to investigate the effect of nimesulide on the small intestine mucositis induced by methotrexate (MTX) in rats. Experimental animals were divided into the control group, MTX group (MTXG) and nimesulide+MTX administered group (NMTXG) with eight rats per group. The control and MTXG groups were given distilled water by gavage and the NMTXG was given nimesulide 100 mg/kg orally. After one hour, the NMTXG and MTXG rat groups were administered oral MTX 5 mg/kg. This procedure was repeated once a day for 15 days and the rats were sacrificed. The duodenum and jejunum of each rat was removed for the assessment of biochemical markers and histopathological evaluation. Malondialdehyde (MDA) and myeloperoxidase (MPO) levels were significantly higher in the duodenal and jejunal tissues of the animals which received MTX, compared to the control and NMTXG (P<0.001). Also, the levels of total glutathione (tGSH), glutathione reductase (GSHRd), glutathione peroxidase (GSHPx), catalase (CAT) and superoxide dismutase (SOD) were significantly lower in the MTXG (P<0.001) compared to other groups. MTX led to villus and crypt epithelial damage and inflammation containing marked PMNL and eosinophils in the intestinal tissues histopathologically. Whereas, there was only mild irregularities in the villus structures of the NMTXG. Nimesulide protected the small intestines against damage by MTX. Intestinal mucositis caused by MTX may be preventable by co-administered nimesulide.

Simultaneous Quantification of Baricitinib and Methotrexate in Rat Plasma by LC-MS/MS: Application to a Pharmacokinetic Study.

Efficacy assessments using a combination of baricitinib and methotrexate necessitate the development of an analytical method for the determination of both drugs in plasma with precision. A high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the simultaneous determination of baricitinib and methotrexate in rat plasma. Extraction of baricitinib, methotrexate, and tolbutamide (internal standard; IS) from 50 µL of rat plasma was carried out by protein precipitation with methanol. Chromatographic separation of the analytes was performed on the YMC pack ODS AM (150 mm × 4.6 mm, 5 µm) column under gradient conditions with methanol: 2.0 mM ammonium acetate buffer as the mobile phases at a flow rate of 1 mL/min. The precursor ion and product ion transition for both analytes and IS were monitored on a triple quadrupole mass spectrometer, operated with selective reaction monitoring in positive ionization mode. The method was validated over a concentration range of 0.5–250.00 ng/mL for baricitinib and methotrexate. Mean extraction recoveries for baricitinib, methotrexate, and IS of 86.8%, 89.4%, and 91.8% were consistent across low, medium, and high QC levels, respectively. Precision and accuracy at low, medium, and high quality control levels were less than 15% across the analytes. Benchtop, wet, freeze-thaw, and long-term stability were evaluated for both of the analytes. The analytical method was applied to support the pharmacokinetic study of simultaneous estimation of baricitinib and methotrexate in Wistar rats. Assay reproducibility was demonstrated by reanalysis of 18 incurred samples

The Ameliorative Effect of Propolis Extract (bee glue) Against Oxidative Damage Induced by Methotrexate in Rats

Methotrexate (MTX) is an anti-folate used to treat cancer and some inflammatory diseases. The efficacy of MTX is often limited by its severe toxicity. Propolis is one of the most significant bee products that has an important role in balancing antioxidant systems and has an anti-peroxidant effect on several tissues. The present study was conducted to investigate the ameliorative effect of propolis against the methotrexate induced oxidative damage on blood, liver and kidney in rats. A total of one hundred and twenty male Wistar albino rats of mean body weights 90 ± 5 g were divided into 5 groups. Rats were administered their respective doses of propolis and/or MTX for 3, 6 and 9 weeks intervals. Control groups (G1 and G2), propolis group (G3), methotrexate group (G4) and methotrexate and propolis co-administered group (G5).The results showed that the MTX significant reduced hemoglobin concentration, hematocrit %, MCHC value, RBCs, WBCs and platelets count and significantly increased MCV and lymphocytes %. Moreover, it caused a significant increase in MDA level and significant decrease in GSH concentration in liver tissue as compared to normal control group (G1) in a time dependent manner. MTX administration also caused significant increase in serum AST, ALT and ALP activities in a time dependent manner, but a significant increase in total bilirubin only in 9 weeks as compared to normal control (G1). On the other hand, MTX impaired kidney function as reflected by a significant increase in serum urea and creatinine levels and decrease in serum uric acid level as compared to normal control group. Results suggest that administration of propolis extract (G5) help reduce the deleterious effects due to MTX injection in a time dependent manner, whereas, administration for 9 weeks offers better alleviation over 6 or 3 weeks. Data showed that long term administration of MTX for 9 weeks produce maximum damage over 6 or 3 weeks. Propolis extract administration also recovered the structural and functional integrity of the hepatic cells.

Effect of growth hormone on testicular dysfunction induced by methotrexate in rats

Introduction & objective: Methotrexate (MTX) is a chemotherapeutic agent that used for the treatment of a variety of tumors and inflammatory diseases. Methotrexate cause defective oogenesis and spermatogenesis, and impair fertility. The aim of this study was planed to see the role of human growth hormone (GH) on testis recovery after treatment with methotrexate in rat. Methods The forty male wistar rat were selected and randomly divided into four groups; control (n=10) and test (n=30). GH groupe was received 0.3mg/kg GH (IP) for 28 consequence day, MTX Group was received 1mg/kg MTX (IP) per week for four consequence week, GH/MTX group was received 0.3mg/kg GH (IP) for 28 consequence day plus 1mg/kg MTX (IP) per week for four consequence week, however the control group just received normal saline (IP). In 14 and 28 days five rats from each group were sacrificed and the testes of rats in whole groups were removed and sperm was collected from epididymis then prepared for analysis. Results: Methodtrexate with 1mg/kg/week dose can cause histopatological and morphological changes in testicular tissue, damage of germinal cells and atrophy of seminiferous tubules. Also cause significant increase of interstitial tissue and capsular thickness and there was significant decrease (p<0.05) in testicular and body weight between this group and others specially control group. Administration of methotrexate cause significant decline in seminiferous tubules diameter and seminiferous epithelium thickness (p<0.05). There were no obvious changes seen in morphmetrical parameters between MTX/GH and control groups. In group that was treated with methodtrexate results shows significant decrease (p<0.05) in number, motility and viability of spermatozoa and testosterone concentration. Administration of GH in addition with methotrexate reduced the effects of methotrexate on sperm parameters and testosterone concentration even completely or partially. Conclusion These results suggested that GH had protective effect on approximately all destructive effects caused by MTX in rat testis and improved sperm parameters.

AB0127 Prophylactic and Therapeutic Administration of β-Glucan from Pleurotus Ostreatus Decreases the Development of Arthritis and Potentiates Methotrexate Treatment in Rats with Adjuvant Arthritis

BackgroundBeta-glucans are naturally occurring polysaccharides, which are integral constituents of the cell wall of bacteria, plants and yeasts. They modify biological response, have anticarcinogenic activity and enhance non-specifically the host...

Abrogation by Ginkgo Byloba Leaf Extract on Hepatic and Renal Toxicity Induced by Methotrexate in Rats

Methotrexate (MTX) is used as a chemotherapeutic agent and its anti-oxidant activity is used to treat many cancer types. The present study aimed to examine the possible modifying effects of Ginkgo biloba leaf extract (GLE) against hepatic and renal toxicity induced by MTX in rats. A total 60 male albino rats were equally divided into six groups; the first and second groups were the control and GLE groups respectively while the 3rd group was MTX rat group; the 4th and 5th groups were Co- and post treated MTX rat with GLE respectively and the 6th group was MTX self treated rat group. Serum GPT, GOT, urea, creatinine, uric acids and MDA levels in MTX group showed a significant increase when compared with control group, in contrast, MTX-treated group also exhibited a significant decrease in liver antioxidant machinery represented by GSH, catalase, SOD and total protein. Administration of GLE combined with MTX improved the liver and kidney damages induced by MTX. Histopathological and evidence, together with observed CD68 immunoreactivity, supported the detrimental effect of MTX and the ameliorating effect of GLE on liver and kidney toxicities. GLE possessed various protective mechanisms against MTX-induced liver and kidney toxicity throughout Co- and post- treatment. We can conclude that Co-treatment with GLE has beneficial properties and can reduce the liver and kidney damages and toxicity induced by MTX.

Preventive Action of Honey on Methotrexate Induced Intestinal Mucositis in Albino Rats (Immunohistochemical Study)

Background and Objectives: Gastrointestinal mucositis is a severe and debilitating side effect of chemotherapy. Methotrexate (MTX) represents one of the most potent classical anti-tumor drugs. The aim of this study was to determine the preventive effects of honey on the intestinal mucositis caused by methotrexate in rats. Materials and Methods: In current study forty females Albino rats, weighing 250-300g were used in the study. For the induction of mucositis, 60 mg/kg of MTX was administered intraperitoneally to each animal in the study group on day 4. The control animals were intraperitoneally injected by normal saline in the same manner and dose like MTX. At the beginning of the experiment, the rats in each group were randomly divided into two groups: Distilled water treated group and honey treated group (10 animals each). A volume of distilled water equal to honey was given by intragastric gavage tube, while the other group was gavaged with honey at a dose of (2.5g/kg) two times daily. The body weight for each rat was measured and then the animals were sacrificed on day 8. In each experiment, one centimeter of prox¬imal jejunum was removed for histopathological, intestinal morphometry, and immunohistochemical analysis using Ki-67 and Bcl-2 immunolabeling. Results: The MTX / honey treated group showed significantly (p 0.05) were seen in the Ki-67 and Bcl-2 immune expression in comparison with the MTX/water treated group. Conclusion: Honey at a dose of 2.5g/kg two times daily, produced a partial protection against methotrexate induced mucositis in rat by decreasing the histopathological scores.

Effects of Streptococcus thermophilus TH-4 in a rat model of doxorubicin-induced mucositis

Background.Mucositis is a debilitating intestinal side effect of chemotherapeutic regimens. Probiotics have been considered a possible preventative treatment for mucositis. Streptococcus thermophilus TH-4 (TH-4), a newly identified probiotic, has been shown to partially alleviate mucositis induced by administration of the antimetabolite chemotherapy drug, methotrexate in rats; likely mediated through a mechanism of folate production. However, its effects against other classes of chemotherapy drug have yet to be determined. Aims.The authors investigated the effects of TH-4 in a rat model of mucositis induced by the anthracycline chemotherapy drug, doxorubicin. Methods. Gastrointestinal damage was induced in female Dark Agouti rats (148.3 ± 1.5 g) by intraperitoneal injection of doxorubicin (20 mg/kg). Animals recieved a daily oral gavage of TH-4 at 109 cfu/ml or skim milk (vehicle) from days 0 to 8. At day 6, rats were injected with either saline or doxorubicin. At kill, small intestinal tissues were collected for determination of sucrase and myeloperoxidase (MPO) activities and histological assessment. Results. Body weight was significantly decreased by doxorubicin compared with normal controls (p < 0.05). Histological parameters, such as crypt depth and villus height, were also significantly decreased by doxorubicin. TH-4 partially prevented the loss of body weight induced by doxorubicin (2.3% compared with 4%), but provided no further therapeutic benefit. Conclusions.The minimal amelioration of doxorubicin-induced mucositis by TH-4 further supports folate production as a likely mechanism of TH-4 action against methotrexate-induced mucositis. Further studies into TH-4 are required to confirm its applicability to other conventional chemotherapy regimens.

Noni increased the systemic exposure of methotrexate in rats through inhibition on multi-drug resistance protein 2 (MRP 2) and breast cancer resistance protein (BCRP)

Methotrexate (MTX) is an important immunosuppressant with narrow therapeutic window. The transport of MTX was associated with multi-drug resistance proteins (MRPs) and breast cancer resistance protein (BCRP). Noni fruit is a phenolics rich health food. This study set out to investigate the effect of noni juice on MTX pharmacokinetics and the underlying mechanisms. Rats were administered MTX with and without single dose or the 7th dose of noni juice (6mL/kg). The serum MTX concentrations were determined using a specific monoclonal fluorescence polarization immunoassay. The results revealed that single dose and multiple doses of noni juice significantly increased the AUC0–t, Cmax and MRT of MTX. A further inquiry into the underlying mechanism indicated that noni metabolites inhibited the activities of MRP 2 and BCRP. In conclusion, concomitant intake of noni juice significantly increased the systemic exposure of MTX through inhibition on MRP 2 and BCRP.

Enhanced antitumor efficacy by methotrexate conjugated Pluronic mixed micelles against KBv multidrug resistant cancer

A methotrexate (MTX) conjugated polymeric mixed micelles for MDR cancer therapy was developed in this study. To the best of our knowledge, MTX was firstly reported to be conjugated with Pluronic P105 (P105-MTX). The Pluronic F127 and P105-MTX polymeric mixed micelles (F127/P105-MTX) were fabricated by thin-film hydration technique, and performed superiority over physically entrapped MTX mixed micelles in drug loading capacity. The drug loading of MTX in F127/P105-MTX was found to be 3.42-fold higher than that of physically entrapped MTX mixed micelles. By conjugated to Pluronic, the amount of MTX in mixed micelles was increased 3.42-fold. In vitro cytotoxicity, cell apoptosis and cell cycle arrest studies also demonstrated that F127/P105-MTX had better antitumor efficacy in KBv MDR cells compared to that of physically entrapped mixed micelles. In comparison with MTX injection, F127/P105-MTX can significantly enhance blood circulation time of MTX in rats. Moreover, a much stronger antitumor efficacy in KBv xenografts mice was observed in F127/P105-MTX group than that of MTX. Therefore, MTX-conjugated mixed micelles might be an effective platform for delivering chemotherapeutic agents to MDR tumors.

Effect of Mirtazapine on Gastric Oxidative Stress and DNA Injury Created With Methotrexate in Rats

Effect of Mirtazapine on Gastric Oxidative Stress and DNA Injury Created With Methotrexate in Rats

St. John's wort significantly increased the systemic exposure and toxicity of methotrexate in rats

St. John's wort (SJW, Hypericum perforatum) is one of the popular nutraceuticals for treating depression. Methotrexate (MTX) is an immunosuppressant with narrow therapeutic window. This study investigated the effect of SJW on MTX pharmacokinetics in rats.Rats were orally given MTX alone and coadministered with 300 and 150mg/kg of SJW, and 25mg/kg of diclofenac, respectively. Blood was withdrawn at specific time points and serum MTX concentrations were assayed by a specific monoclonal fluorescence polarization immunoassay method.The results showed that 300mg/kg of SJW significantly increased the AUC0−t and Cmax of MTX by 163% and 60%, respectively, and 150mg/kg of SJW significantly increased the AUC0−t of MTX by 55%. In addition, diclofenac enhanced the Cmax of MTX by 110%. The mortality of rats treated with SJW was higher than that of controls.In conclusion, coadministration of SJW significantly increased the systemic exposure and toxicity of MTX. The combined use of MTX with SJW would need to be with caution.

Induction of hyperchromic microcytic anaemia by repeated oral administration of methotrexate in rats

Anaemia is a significant prognostic factor in cancer patients receiving anticancer drugs such as methotrexate (MTX). This study focuses on the effects of toxicological changes on the hematopoietic systems in male and female Wistar Hannover rats when MTX is orally administered at a dose of 0, 0.05, 0.15, or 0.45 mg/(kg·day) for a period of 28 days. Both male and female rats receiving 0.45 mg/kg MTX showed a decrease in the haemoglobin concentration (Hb), haematocrit, and erythrocyte count. Female rats showed a decrease in mean corpuscular volume (MCV) and an increase in cell mean Hb (CHCM) in total erythrocytes, including the mature erythrocytes. These results indicate that MTX causes the production of small, mature erythrocytes that contain a high concentration of Hb. MTX reduced the number of peripheral reticulocytes but produced the cells with a large size and a high concentration of Hb, as demonstrated by the reticulocyte MCV and CHCM as well as the content of haemoglobin per reticulocyte (CHr). Consistent with these findings, bone marrow haematopoiesis was impaired by MTX, as there was a reduction in erythroid count in rats of both sexes. The number of cells of the myeloid lineage reduced in female rats, followed by a reduction in the total leukocyte and neutrophil counts in peripheral blood. Thrombocytopenia was detected in a small population of rats. These results indicate that MTX induces hyperchromic microcytic anaemia and pancytopenia, and the use of MCV and CHCM in mature erythrocytes and reticulocytes, along with the CHr, gives a better understanding of the development and nature of anaemia.

Hepatoprotective Effect of Taurine Against Oxidative Stress Due to Methotrexate in Rat

Problem statement: Taurine, 2-aminoethanesulphonic acid is an essential amino acid. It is present at high concentrations in many tissues. It plays important roles in numerous physiological functions including conjugation with bile acids, modulation of calcium levels and maintenance of osmolarity, antioxidation and stabilization of membranes. Methotrexate (MTX), as the most disease modifying anti-rheumatic drug used for Rheumatoid Arthritis (RA), has been available for clinical use since 1951. The main scope of this study was to assessment of probable Hepatoprotective effect of taurine as it antioxidant action against oxidative stress due to methotrexate in rats. Approach: Eighty male Wistar rats, weighing 200±20 gr, were obtained from the animal breeding center of Islamic Azad University. The rats were divided into 4 equal groups of 20 animals including: 1-normal control, 2-normal rats treated with taurine, 3- taurine + methotrexate and 4-methotrexate. Results: Methotrexate-induced hepatotoxicity as reflected by elevated serum ALT, AST and GGT levels (p<0.001) and reduced serum total protein level (p<0.001). Conclusion: Taurine has a protective action against hepatotoxins and suggest that taurine may find clinical application against a variety of toxins where cellular damage is a consequence of reactive oxygen species.

Injectable thermosensitive hydrogels for intra‐articular delivery of methotrexate

AbstractIntra‐articular drug delivery is the preferred approach for targeting pharmacologic treatment directly at the joints to reduce undesirable side effects associated with systemic drug delivery. In this study, a controlled delivery system of methotrexate (MTX) based on injectable thermosensitive poly(ε‐caprolactone)‐poly(ethylene glycol)‐poly(ε‐caprolactone) (PCL‐PEG‐PCL, PEP) hydrogels was developed for the intra‐articular drug delivery. The thermosensitive PEP copolymers were prepared by ring‐opening polymerization. The synthesized PEP copolymers were characterized for their structure, composition, and the sol‐to‐gel transition. The in vitro MTX release from the PEP hydrogels was studied. MTX plasma concentration following intra‐articular injection into healthy rats was determined by HPLC. Biocompatibility was confirmed by histology analysis after the intra‐articular injection. The synthesized PEP copolymers aqueous solutions formed in situ gel rapidly after the injection. PEP hydrogels showed the ability to control the release of incorporated MTX. Following intra‐articular injection, the PEP hydrogels decreased the clearance rate of MTX in the joint cavity. The maximum plasma concentrations of MTX in rats injected with free MTX were threefold higher than that of the groups injected with MTX hydrogels. These results suggest that the intra‐articular delivery of the PEP hydrogels may be a viable strategy for the controlled release of drugs for treating arthritis diseases. © 2011 Wiley Periodicals, Inc. J Appl Polym Sci, 2011

Monoketocholate can decrease transcellular permeation of methotrexate across Caco-2 cell monolayers and reduce its intestinal absorption in rat

Abstract Objectives Bile salts have been shown to decrease the absorption of methotrexate in the rat intestine by an unknown mechanism. We aimed to examine this effect. Methods We assessed apical-to-basolateral (AP-BL) permeation of methotrexate (5 μM) across Caco-2 cell monolayers pretreated with various concentrations (0, 0.25, 0.5, 1, 3 and 5 mm) of sodium cholate or its semisynthetic analogue, sodium 12-monoketocholate. We also determined the effect of orally administered 12-monoketocholate on the intestinal absorption of methotrexate in rats to evaluate a possible in-vitro–in-vivo correlation. Key findings It was found that sodium cholate and sodium 12-monoketocholate decreased the AP-BL permeation of methotrexate at low concentrations (maximal inhibition at 0.25 and 1 mm, respectively) and increased it at higher concentrations. Determination of [14C] mannitol permeation and electrical resistance of monolayers during experiments showed that membrane integrity was not compromised at low concentrations of bile salts but was disrupted at higher concentrations. Subsequently, we examined the effect of the simultaneous oral administration of sodium 12-monoketocholate (4, 20, 40 and 80 mg/kg) on the intestinal absorption of methotrexate in rats after an oral dose (5 mg/kg). The pharmacokinetic study showed that 12-monoketocholate at 4 and 20 mg/kg did not change the methotrexate area under the serum concentration–time curve whereas sodium 12-monoketocholate at 40 and 80 mg/kg significantly reduced it. Conclusions Sodium 12-monoketocholate appears to decrease the intestinal absorption of methotrexate in rats by inhibition of transcellular active transport.

Site-specific contribution of proton-coupled folate transporter/haem carrier protein 1 in the intestinal absorption of methotrexate in rats

Abstract Objectives Methotrexate is reportedly a substrate for proton-coupled folate transporter/haem carrier protein 1 (PCFT/HCP1) and reduced folate carrier 1 (RFC1). In this study, we examined the contribution of PCFT/HCP1 and RFC1 in the intestinal absorption of methotrexate in rats. Methods Western blot analysis was carried out to evaluate the protein levels of PCFT/HCP1 and multidrug resistance-associated protein 2 in brush-border membrane of rat small intestine. Mucosal uptake of methotrexate was studied in the rat everted small intestine and an in-situ intestinal perfusion study of methotrexate was also carried out in rats. Key findings In transport studies using everted intestine, the mucosal methotrexate influx rate in proximal intestine at pH 5.5 was significantly greater than that at pH 7.4. Coadministration of folate or its analogues, such as folinate and 5-methyltetrahydrofolate, substrates for both PCFT/HCP1 and RFC1, significantly suppressed the methotrexate influx at pH 5.5, whereas thiamine pyrophosphate, an inhibitor for RFC1 alone, exerted no significant effect. Western blot analysis showed higher PCFT/HCP1 expression in proximal than distal small intestine. In distal small intestine, methotrexate influx rate was low and was not pH dependent. Also, folate and its analogues exerted no significant effect on methotrexate absorption. Conclusions Based on the present and our previous results, the site-specific contributions of various transporters including PCFT/HCP1 in methotrexate intestinal absorption were discussed. The variation in luminal pH and the involvement of multiple transporters in methotrexate absorption may cause variation in oral bioavailability among patients.

Inhibition of hippocampal cell proliferation by methotrexate in rats is not potentiated by the presence of a tumor

Methotrexate is a widely used cytostatic in chemotherapy cocktails for the treatment of cancer but is associated with cognitive impairment. Previous animal studies indicated that methorexate decreases hippocampal cell proliferation, which might contribute to the observed cognitive impairment. However, clinical studies have shown that cognitive impairment can also be noticed in some cancer patients before any systemic treatment is initiated. We aim in the present study to discern whether hippocampal cell proliferation is negatively affected by tumor growth and if the presence of a tumor amplifies the effects of methotrexate. Buffalo rats were subcutaneously injected with PBS or Morris Hepatoma 7777 cells to induce a tumor. Two weeks after this injection the animals received an intraperitoneal injection of methotrexate or saline. Three weeks later hippocampal cell proliferation was quantified using immunohistochemical staining. Treatment with Morris Hepatoma 7777 cells decreased the number of proliferating cells as compared to control animals. An overall tumor effect was absent mainly because methotrexate treatment significantly decreased cell proliferation with no differences between animals with or without a tumor. Neither methotrexate nor the tumor induced pica behavior. These findings indicate that although the presence of a tumor reduces hippocampal cell proliferation it does not affect the negative effect of methotrexate on this plasticity marker. Since sickness behavior is not induced by methotrexate or tumor presence it does not play a role in the development of cognitive deficits. This study further indicates that the effects of methotrexate on brain and behavior can be studied in healthy animals.