Przemysław Ł. Mikołajczak

The influence of mianserin on TNF-α, IL-6 and IL-10 serum levels in rats under chronic mild stress

BACKGROUND Antidepressants are known to affect the immunological system through mechanisms which are not completely understood. The aim of the present study was to evaluate the effect of the atypical antidepressant mianserin on the levels of tumor necrosis factor alpha (TNFα), interleukin-6 (IL-6) and interleukin-10 (IL-10) in the blood of rats in an experimental model of depression. METHODS Male Wistar rats were subjected to chronic mild stress (CMS) according to Willners method for 6 weeks. Following the development of anhedonia, the stressed and control rats (non-stressed animals) were treated with mianserin (10 mg/kg ip, twice daily) for three weeks. On the last day of the experiment, a lipopolysaccharide (LPS, 100 μg/kg ip) was injected to mianserin- or vehicle-treated rats. TNFα, IL-6 and IL-10 levels in the blood of the rats were assayed using ELISA methods. RESULTS The results indicated a significantly increased TNFα level in stressed animals when compared with the non-stressed (control) group. The levels of IL-6 and IL-10 were also elevated, especially after LPS administration. Treatment with mianserin resulted in a significant lowering of TNFα and IL-6 levels both in LPS-treated and LPS-untreated animals. There was also a decrease in IL-10 concentration in LPS-treated stressed animals. CONCLUSIONS The results confirm an increase in proinflammatory cytokines in the blood of rats with experimentally induced depression and show the protective role of the activity of mianserin on the cytokine levels, expressed in a lowering of TNFα and IL-6 levels in stressed animals, and of IL-10 levels after LPS administration.

Evaluation of anti-inflammatory and analgesic activities of extracts from herb of Chelidonium majus L.

The aim of the study was to evaluate analgesic activity (“hot plate” test), anti-inflammatory activity (carrageenan-induced paw edema) and locomotor activity in rats under the influence of three fractions of Chelidonium majus herb extract: full water extract (FWE), protein enriched fraction (PEF), and non-protein fraction (NPF). Effects of the fractions on the level of chosen cytokines and their mRNA levels were also assessed using lipopolysaccharide (LPS) administration as a proinflammatory cue. All fractions and diclofenac did not affect the locomotor activity of rats in comparison with the control group. FWE and PEF three hours after administration showed statistically significant analgesic activities comparable to morphine (p < 0.05). A slight reduction in rat paw edema was observed after three (comparable with diclofenac) and six hours in the NPF group. FWE revealed a statistically significant pro-inflammatory effect after three hours in comparison with the control group. Peripheral IL-1 and IL-4 cytokine concentrations were reduced under FWE and NPF, PEF fractions. The combination of FWE, PEF and NPF together with LPS showed only the effects of LPS. We suggest that protein enriched fraction (PEF) produced centrally mediated (morphine-like) analgesic action, whereas the anti-inflammatory potential was shown only after LPS-induced inflammation. The precise mechanisms involved in the production of anti-nociceptive and anti-inflammatory responses of studied fractions are not completely understood, but they may be caused rather by the presence of protein more than alkaloids-enriched fraction. This fraction of the extract could be used as an alternative therapy for the prevention of inflammatory-related diseases in the future, but further studies are needed.

Impact of Panax ginseng and Ginkgo biloba extracts on expression level of transcriptional factors and xenobiotic-metabolizing cytochrome P450 enzymes

Summary Introduction: Despite widespread use of Panax ginseng and Ginkgo biloba, the data on the safety as well as herb-drug interactions are very limited. Therefore, we postulate that P. ginseng and G. biloba may modulate the activity and content of cytochrome P450 isozymes involved in the biotransformation of diverse xenobiotic substances. Objective: The aim of our study was to determine the influence of herbal remedies on the expression level of CYP enzymes and transcriptional factors. Methods: Male Wistar rats were given standardized Panax ginseng (30 mg/kg p.o.) or standardized Ginkgo biloba (200 mg/kg p.o.) for 3 and 10 days. The expression in liver tissue was analyzed by realtime PCR method. Results: Our results showed a decrease of CYP3A1 (homologue to human CYP3A4) mRNA level after P. ginseng extract treatment. The CYP2C6 (homologue to human CYP2C9) expression was also reduced. Additionally, after 10 days of the treatment with P. ginseng an increase of CYP1A1 (homologue to human CYP1A1) and CYP1A2 (homologue to human CYP1A2) expression was observed. Moreover, G. biloba extract also caused an increase of expression level for CYP1A1, CYP2C6, CYP3A1 and CYP3A2. Conclusion: Our findings suggest that herbal extracts can modulate the expression of transcriptional factors and CYP enzymes involved in xenobiotic metabolism and chemical carcinogenesis. Streszczenie Wstęp: Mimo powszechnego stosowania Panax ginseng i Ginkgo biloba dane dotyczące bezpieczeństwa, a także interakcji pomiędzy preparatami roślinnymi a lekami syntetycznymi są bardzo ograniczone. W niniejszych badaniach założono, iż żeń-szeń oraz miłorząb mogą modulować aktywność i zawartość izoenzymów cytochromu P450 biorących udział w biotransformacji różnych substancji ksenobiotycznych. Cel: Określenie wpływu preparatów roślinnych na poziom ekspresji enzymów CYP i ich czynników transkrypcyjnych. Metody: Szczurom rasy Wistar podawano standaryzowany Panax ginseng (30 mg/kg) oraz Ginkgo biloba (200 mg/kg) przez 3 do 10 dni. Ekspresję w tkance wątrobowej analizowano za pomocą metody PCR w czasie rzeczywistym. Wyniki: Uzyskane wyniki wykazały spadek poziomu mRNA CYP3A1 (homolog ludzkiego enzymu CYP3A4) po podaniu ekstraktu z żeń-szenia. Ekspresja genu CYP2C6 (homolog ludzkiego enzymu CYP2C9) również uległa obniżeniu. Dodatkowo, obserwowaliśmy wzrost ekspresji CYP1A1 (homolog ludzkiego enzymu CYP1A1) i CYP1A2 (homolog ludzkiego enzymu CYP1A2) po 10 dniach stosowania P. ginseng. Ponadto, ekstrakt z G. biloba spowodował również wzrost poziomu mRNA CYP1A1, CYP2C6, CYP3A1 i CYP3A2 w modelu in vivo. Wnioski: Badania sugerują, że wyciągi roślinne mogą modulować ekspresję czynników transkrypcyjnych i enzymów CYP uczestniczących w metabolizmie ksenobiotyków i chemicznej karcynogenezie.

Influence of salidroside, a neuroactive compound of Rhodiola rosea L., on alcohol tolerance development in rats

Summary Introduction: In recent years, the search for potential neuroprotective properties of salidroside and its ability to influence the activity of nervous system become the subject of intense studies of many research groups. None of these studies, however, include an attempt to determine the effect of salidroside on the course of alcohol tolerance in vivo. Objective: The aim of this study was to investigate the ability of salidroside to inhibit the development of alcohol tolerance in rats, determining whether the effect of its action may occur in a dose-dependent manner, reducing both metabolic and central tolerance without affecting body temperature in control rats. Methods: Male Wistar rats were injected daily with ethanol at a dose of 3 g/kg for 9 consecutive days to produce ethanol tolerance. Salidroside in two doses (4.5 mg/kg and 45 mg/kg b.w.) or vehiculum was administered orally. On the 1st, 3rd, 5th and 8th day a hypothermic effect of ethanol was measured, while the loss of righting reflex procedure was performed on the 2nd, 4th, 6th and 7th day. On the 9th day rats were treated with salidroside, sacrificed 1 h after ethanol injections and blood was collected for blood-ethanol concentration measurement. Results: Salidroside at a dose of 45 mg/kg inhibited the development of tolerance to hypothermic and sedative effects of ethanol, whereas insignificant elevation of blood-ethanol concentration was observed. The dose of 4.5 mg/kg b.w. had minimal effect, only small inhibition of tolerance to hypothermic action was observed. Salidroside affected neither body mass growth nor body temperature in non-alcoholic (control) rats. Conclusions: Results of the study indicate that salidroside at a dose of 45 mg/kg inhibited the development of tolerance to the hypothermic effect of ethanol. Observed inhibition of tolerance to the sedative effect of ethanol seems to be associated with salidroside influence on the central nervous system. A comprehensive explanation of the abovementioned observations requires further pharmacological and pharmacodynamic studies.

Diet based on oil of seeds of Brassica napus. Implications for the prevention and treatment of prostate diseases

Summary In last centuries, human diet has been modified due to changes in the technological, economic, and cultural mode of the social life. The rapid development of the technology and advances in food production and processing of plants resulted in changes in the structure of consumption. The consequence is an growing interest among researchers, but also among consumers in searching for new alternatives to classical pharmacology, preventive solutions, and therapeutic strategies of age related, chronic diseases and cancer. In this paper, the authors have attempted to assess the benefits of usage in diet cold-pressed oils extracted from the seeds of Brassica napus (oilseed rape) with particular emphasis on the evidence of possible use of B. napus oil, and other bio-active substances contained therein, in the prevention and symptomatic treatment of prostate diseases, including benign prostatic hyperplasia (BPH). The authors postulate that the information presented in this paper may be important and helpful in the development of new, effective and safe forms of prevention and treatment of progressive age-related disorders (phytotherapeutics) and nutritional strategies (dietary supplements) based on seeds oil of Brassicace family representatives, especially Brassica napus (pollen). Streszczenie W ostatnich stuleciach ludzka dieta została istotnie zmodyfikowana w wyniku zmian trybu technologicznych, ekonomicznych i kulturalnych. Szybki rozwój technologii i postęp w produkcji i przetwórstwie żywności z roślin spowodował zmiany w strukturze konsumpcji. Konsekwencją tych zdarzeń jest wzrost zainteresowania naukowców oraz konsumentów poszukiwaniem nowych, alternatywnych strategii żywieniowych, a także profilaktyką schorzeń towarzyszących okresowi starzenia się, jak również chorób przewlekłych i nowotworowych. W niniejszej pracy autorzy podjęli próbę przybliżenia korzyści wynikających ze stosowania diety zawierającej tłoczony na zimno olej uzyskiwany z nasion Brassica napus (rzepak), ze szczególnym uwzględnieniem dowodów świadczących o możliwości wykorzystania oleju z B. napus, substancji bioaktywnych w nim zawartych w prewencji i leczeniu objawowym schorzeń prostaty, w tym BPH. Autorzy sugerują, że informacje zawarte w niniejszej pracy mogą być istotne i pomocne w opracowaniu nowych, skutecznych i bezpiecznych form prewencji i leczenia objawowego schorzeń postępujących wraz z wiekiem (fitoterapeutyki) oraz strategii żywieniowych (suplementy diety) opartych na bazie oleju z nasion wybranych przedstawicieli rodziny Brassicaceae, w tym Brassica napus.

SRC kinase mRNA transcription changes in testosterone-induced rat ventral prostate lobes under the influence of Epilobium angustifolium extract

Abstract The aim of this study was to investigate the influence of standardized crude aqueous Epilobium angustifolium L. extract [100 mg/kg/day, p.o.] on the expression level of SRC kinase mRNA - a representatives of non-genomics xenobiotics signaling pathway in prostate ventral lobes of testosterone-induced, castrated rats. We have shown that in all analyzed groups induced by testosterone an elevation of SRC kinase mRNA transcription was observed, in comparison to control animals (not receiving the testosterone), (p<0.05). Finasteride in rats induced by testosterone caused the strongest inhibition of SRC mRNA transcription (p<0.05). In rats receiving testosterone and the plant extract a ca. 90% decrease of mRNA level was observed vs. testosterone-induced animals (p<0.05), while in testosterone-induced animals receiving concomitantly E. angustifolium extract and finasteride the observed reduction reached 87.3% (p<0.05). We did not observed, however, any positive feedback between studied plant extract and finasteride in the inhibitory activity (p<0.05). Further experimental studies should be performed in order to the understanding the molecular basis of interactions, the efficacy and safety of tested plant extract. Streszczenie Celem pracy było zbadanie wpływu standaryzowanego ekstraktu z ziela Epilobium angustifolium L. [100 mg/kg/dzień, p.o.] na poziom transkrypcji mRNA kinazy SRC - przedstawiciela androgenozależnego, niegenomowego szlaku sygnalizacji komórkowej w brzusznym płacie prostat indukowanych hormonalnie, kastrowanych szczurów. We wszystkich grupach szczurów indukowanych testosteronem zaobserowaliśmy wzrost liczby transkryptów kinazy SRC, w porównaniu do zwierząt kontrolnych (p<0.05). U szczurów indukowanych hormonalnie finasteryd wykazał najsilniejsze właściwości hamujące transkrypcję mRNA (p<0.05). U zwierząt otrzymujących testosteron wraz z wyciągiem z E. angutifolium stwierdziliśmy obniżenie liczby badanych transkryptów o blisko 90%, w porownaniu ze zwierzętami z grupy kontrolnej, indukowanymi testosteronem (p<0.05). U szczurów otrzymujących jednocześnie testosteron i ekstrakt roślinny łącznie z finasterydem redukcja poziomu mRNA kinazy SRC sięgała 87.3% (p<0.05). Nie zaobserwowaliśmy dodatniego sprzężenia w inhibicji transkrypcji badanego mRNA u szczurów otrzymujących badany ekstrakt wraz z finasterydem (p<0.05). Istnieje potrzeba przeprowadzenia dalszych badań zmierzających do wyjaśnienia molekularnego podłoża interakcji, mechanizmu działania oraz skuteczności badanego wciągu roślinnego.

Influence of epilobium angustifolium extract on 5α-reductase type 2 and mapk3 kinase gene expression in rats prostates

Abstract The aim of this study was to investigate the influence of standardized Epilobium angustifolium L. extract [100 mg/kg/day, p.o.] on the expression level of 5α-reductase type 2 (Srd5ar2) mRNA and Mapk3 mRNA a representative of non-genomic xenobiotics signaling pathway. It was shown that plant extract from the E. angustifolium showed a slight tendency to reduce prostate weight in hormonally induced animals (p>0.05) and in testosterone induced animals receiving both, extract and finasteride (p<0.05). Finasteride in rats induced by testosterone caused a smaller decrease in the level of mRNA 5α-steroid reductase 2 (SRd5ar2), than in rats treated with the hormone and studied plant extracts. In general, an increase in the amount of MAPK3 mRNAs in testosterone-induced groups of rats receiving tested plant extract with or without finasteride was observed, while the expression of type 2 5α-steroid reductase decreased (p<0.05). Further experimental studies should be performed in order to understand the molecular basis of interactions, the efficacy and safety of tested plant extracts. Streszczenie Celem pracy było zbadanie wpływu standaryzowanego ekstraktu z ziela Epilobium angustifolium L. [100 mg/kg/dzień, p.o.] na poziom ekspresji mRNA 5α-reduktazy typu 2 (SRd5ar2) oraz mRNA kinazy MAPK3 - przedstawiciela androgenozależnego, nie-genomowego szlaku sygnalizacji komórkowej. W zastosowanym modelu eksperymentalnym wyciąg z z E. angustifolium wykazał statystycznie nieistotną, niewielką tendencję do zmniejszania masy prostat u zwierząt indukowanych hormonalnie (p>0,05) oraz u szczurów indukowanych testosteronem, otrzymujących zarówno ekstrakt, jak i finasteryd (p<0,05). Finasteryd u szczurów otrzymujących testosteron spowodował mniejsze, aniżeli zakładano, obniżenie poziomu mRNA 5α-reduktazy typu 2 (SRd5ar2), niż u szczurów, którym podano hormon i badany wyciąg (p <0.05). Stwierdziliśmy ponadto, zwiększenie ilości mRNA kinazy MAPK3 u szczurów indukowanych testosteronem otrzymujących badany ekstrakt, wraz z finasterydem lub bez niego, podczas gdy ekspresja reduktazy w tych grupach uległa zwiększeniu (p <0,05). Należy przeprowadzić dalsze badania eksperymentalne w celu zrozumienia molekularnych podstaw oddziaływań, skuteczności i bezpieczeństwa badanych ekstraktów roślinnych.

Effect of Camellia sinensis extract on the expression level of transcription factors and cytochrome P450 genes coding phase I drug-metabolizing enzymes

Abstract Green tea (Camellia sinensis) is widely used as a popular beverage and dietary supplement that can significantly reduce the risk of many diseases. Despite the widespread use of green tea, the data regarding the safety as well as herb-drug interactions are limited. Therefore, the aim of our study was to assess the influence of standardized green tea extract (GTE) containing 61% catechins and 0.1% caffeine on the expression level of rat CYP genes and the corresponding transcription factors expression by realtime PCR. The findings showed that GTE resulted in a significant decrease of CYP2C6 expression level by 68% (p<0.001). In case of CYP3A1 and CYP3A2, the mRNA levels were also reduced by extract but in a lesser degree compared to CYP2C6. Simultaneously the significant increase in the mRNA level of CAR, RXR and GR factors was observed by 54% (p<0.05), 79% (p<0.001) and 23% (p<0.05), respectively after 10 days of green tea extract administration. In addition, there was noted a small increase of CYP1A1 expression level by 21% (p>0.05) was noted. No statistically significant differences were observed for CYP1A2 and CYP2D1/2. In the same study we observed an increase in amount of ARNT gene transcript by 27% (p<0.05) in the long-term use. However, green tea extract showed the ability to stimulate HNF-1α both after 3 and 10 days of treatment by 30% (p<0.05) and 80% (p<0.001), respectively. In contrast, no change was observed in the concentration of HNF-4α cDNA. These results suggest that GTE may change the expression of CYP enzymes, especially CYP2C6 (homologue to human CYP2C9) and may participate in clinically significant interactions with drugs metabolized by these enzymes. Streszczenie Zielona herbata (Camellia sinensis) jest powszechnie stosowana jako napój i suplement diety i może istotnie zmniejszać ryzyko wystąpienia wielu chorób. Pomimo powszechnego zastosowania zielonej herbaty, dane dotyczące bezpieczeństwa jak i interakcji preparatu roślinnego i leku syntetycznego są bardzo ograniczone. Celem badania była ocena wpływu standaryzowanego ekstraktu z zielonej herbaty (GTE) zawierającego 61% katechin i 0,1% kofeiny na poziom ekspresji szczurzych genów CYP i czynników transkrypcyjnych stosując technikę real-time PCR. Wyniki wykazały, że GTE znacznie obniża poziom ekspresji CYP2C6 o 68% (p<0,001). W przypadku CYP3A1 i CYP3A2 poziom mRNA tych genów był również redukowany przez ekstrakt, ale w mniejszym stopniu w porównaniu do CYP2C6. Istotny wzrost w poziomie mRNA obserwowano dla czynników CAR, RXR i GR odpowiednio o 54% (p<0,05), 79% (p<0,001) i 23% (p<0,05) po 10 dniach stosowania ekstraktu. Dodatkowo, zanotowano niewielki wzrost poziomu ekspresji CYP1A1 o 21% (p>0,05). Brak istotnych różnic zaobserwowano dla CYP1A2 i CYP2D1/2. W badaniu wykazano również wzrost ilości transkryptu genu ARNT o 27% (p<0,05) podczas dłuższego stosowania. Ponadto, ekstrakt z zielonej herbaty wykazał zdolność do stymulacji HNF-1α zarówno po 3, jak i 10 dniach trwania eksperymentu odpowiednio o 30% (p<0,05) i 80% (p<0,001). Brak zmian obserwowano w przypadku stężenia cDNA dla HNF-4α. Wyniki te sugerują, że GTE może zmieniać ekspresję enzymów CYP, szczególnie w przypadku CYP2C6 (homolog ludzki CYP2C9) i może uczestniczyć w klinicznie istotnych reakcjach z lekami metabolizowanymi przez te enzymy.

283 TRAMADOL, DIHYDROCODEINE AND METABOLITES LEVELS IN CANCER PATIENTS TREATED WITH CONTROLLED RELEASE DIHYDROCODEINE AND TRAMADOL

that results in the loss of drug effect or requires escalating doses to produce pain relief. It has been shown that chronic administration of morphine induced tolerance to the analgesic effect and apoptosis in central nervous system. The aim of this study was to evaluate the effects of intracerebroventricular (icv) administration of riluzole (anti-glutamatergic drug) on morphine induced tolerance and apoptosis in lumbar spinal cord of rat. Methods: Different groups of rats received morphine (10mg/kg, ip) + vehicle of riluzole [Tween 80–1%] (10ml/rat, icv) or morphine (10mg/kg, ip) + riluzole (20, 40, 80mg/10ml/rat, icv) or riluzole (80mg/10ml/rat, icv) alone, once per day. Nociception was assessed using hot-plate apparatus. Apoptosis was assessed by in situ TdTmediated dUTP-biotin nick end-labeling (TUNEL) method. Bcl2 as anti-apoptotic factor and caspase 3 as proapoptotic agent were evaluated using immunoblotting method. Results: Our findings showed that, riluzole (icv) attenuated morphine tolerance and number of apoptotic cells. Also semiquantitive results showed that the amount of bcl2 was greater when morphine co-administrated with riluzole than morphine only but there are not significant changes in amount of caspase3 between morphine or morphine + riluzole groups. Conclusion: Intracerebroventricular administration of riluzole could attenuate the morphine tolerance and number of apoptotic cells and increased the amount of Bcl2, anti-apoptotic factor, in lumbar spinal cord of rat.