اثرات وابسته به دوز نئوتام بر بیان mRNA ژن‌های کبدی Th و Cyp1a2، سطح دوپامین و هیستوپاتولوژی کبد در موش‌های صحرایی نر

نوع مقاله : مقاله پژوهشی

نویسندگان

1 دانشکده علوم، دانشگاه القاسم الخضراء، 51013، بابل، عراق

2 دانشکده علوم، دانشگاه القاسم الخضراء، 51013، بابل، عراق.

چکیده

هدف: نئوتام یک شیرین‌کننده دی‌پپتیدی با شدت شیرینی بالا و بدون کالری است که از نظر ساختاری با آسپارتام ارتباط دارد و برای استفاده در مواد غذایی و نوشیدنی‌ها تأیید شده است. این مطالعه با هدف بررسی اثرات وابسته به دوز تجویز خوراکی نئوتام بر بیان ژن‌های تیروزین هیدروکسیلاز (TH) و سیتوکروم P450 1A2 (CYP1A2) در کبد، غلظت دوپامین سرم، شاخص‌های عملکرد کبد و ساختار بافتی کبد در موش‌های صحرایی نر بالغ آلبینو طراحی شد.
مواد و روش‌ها: چهار گروه شامل 10 موش صحرایی نر بالغ نژاد ویستار (در مجموع 40 موش) به‌صورت تصادفی انتخاب شدند. یک گروه از حیوانات به‌عنوان گروه کنترل با حلال تیمار شد و سه گروه دیگر به مدت 60 روز متوالی با نئوتام تیمار شدند. دوزهای مورد استفاده شامل 250، 500 یا 750 میلی‌گرم به ازای هر کیلوگرم وزن بدن در روز بود که از طریق گاواژ خوراکی تجویز شد. بیان mRNA ژن‌های کبدی TH و CYP1A2 با استفاده از واکنش زنجیره‌ای پلیمراز کمی بلادرنگ با رونویسی معکوس (qRT-PCR) و با نرمال‌سازی نسبت به ژن مرجع، اندازه‌گیری شد. دوپامین سرم با استفاده از روش ELISA ا ندازه‌گیری شد. عملکرد کبد از طریق اندازه‌گیری ALT، AST، ALP، پروتئین تام و بیلی‌روبین تام ارزیابی شد و مقاطع کبدی نیز از نظر هیستوپاتولوژیک مورد بررسی قرار گرفتند. داده‌ها با استفاده از آزمون ANOVA یک‌طرفه همراه با آزمون‌های پس‌آزمون دانکن و توکی تجزیه‌وتحلیل شدند و ضرایب همبستگی پیرسون بین بیان ژن و شاخص‌های بیوشیمیایی/بافت‌شناختی محاسبه شد.
نتایج: نئوتام باعث کاهش وابسته به دوز بیان mRNA ژن TH در کبد و کاهش همزمان دوپامین سرم شد. در مقابل، افزایش وابسته به دوز بیان mRNA ژن CYP1A2 در کبد و افزایش فعالیت آنزیم‌های ALT و AST مشاهده شد. این تغییرات با افزایش تدریجی شدت التهاب اطراف ورید باب، واکوئله‌شدن سلول‌های کبدی و ایجاد فیبروز خفیف در بالاترین دوز همراه بود. بیان TH با غلظت دوپامین سرم همبستگی مثبت و با بیان CYP1A2 و فعالیت آنزیم‌های کبدی همبستگی منفی داشت.
نتیجه‌گیری: این مطالعه نشان می‌دهد که تغییرات وابسته به دوز در بیان mRNA ژن‌های کبدی Th و Cyp1a2، غلظت دوپامین سرم، شاخص‌های بیوشیمیایی کبد و هیستوپاتولوژی کبد در موش‌های صحرایی نر ممکن است ناشی از قرارگیری مزمن این حیوانات در معرض دوزهای بالای نئوتام باشد. این نتایج بر ضرورت انجام مطالعات بیشتر برای استفاده از ژن‌های مرجع بیشتر، اندازه‌گیری سطوح پروتئینی و بررسی مکانیسم مسیرهای پیام‌رسانی مرتبط تأکید می‌کنند.

کلیدواژه‌ها

عنوان مقاله [English]

Dose-dependent effects of neotame on hepatic Th and Cyp1a2 mRNA expression, dopamine level, and hepatic histopathology in male rats

نویسندگان [English]

  • Randah F. Hassan 1
  • Hassan K. Al-Aawadi 1
  • Rawaa S. A. Al-Azawi 2
1 College of Science, Al-Qasim Green University, 51013, Babylon, Iraq.
2 College of Science, Al-Qasim Green University, 51013, Babylon, Iraq
چکیده [English]

Objective
Neotame is a high-intensity, non-caloric dipeptide sweetener structurally related to aspartame that is approved for use in foods and beverages. Because it is metabolized in the liver and shares structural features with compounds shown to disturb hepatic drug-metabolizing enzymes and catecholaminergic signaling, concerns persist regarding its dose-dependent effects on hepatic gene expression and dopaminergic-related systemic function. This study was designed to evaluate the dose-dependent effects of orally administered neotame on the hepatic expression of tyrosine hydroxylase (TH) and cytochrome P450 1A2 (CYP1A2) genes, serum dopamine concentration, liver function biomarkers, and hepatic histoarchitecture in adult male albino rats.
Material and methods
Four groups of ten adult male Wistar rats (forty rats in total) were randomly selected. One group of animals was a control group treated with solvent and the other three groups were treated with neotame for 60 consecutive days. The doses used were 250, 500, or 750 mg/kg body weight per day, administered by oral gavage. Hepatic TH and CYP1A2 mRNA expression was quantified by quantitative real-time reverse-transcription PCR (qRT-PCR) with normalization to validated reference gene. Serum dopamine was measured by ELISA; liver function was assessed via ALT, AST, ALP, total protein, and total bilirubin; and liver sections were examined histopathologically. Data were analyzed by one-way ANOVA with Duncan's and Tukey's post-hoc test, and Pearson correlation coefficients were calculated between gene expression and biochemical/histological markers.

Results
Neotame produced a dose-dependent downregulation of hepatic TH mRNA and a concomitant reduction in serum dopamine, together with dose-dependent upregulation of hepatic CYP1A2 mRNA and elevation of ALT and AST activities, accompanied by progressively more severe periportal inflammation, hepatocellular vacuolation, and mild fibrosis at the highest dose. TH expression correlated positively with serum dopamine and negatively with CYP1A2 expression and liver enzyme activities.
Conclusion
This study suggests that dose-dependent changes in hepatic Th and Cyp1a2 mRNA expression, serum dopamine concentration, liver biochemical markers, and liver histopathology in male rats may be due to the fact that these animals were chronically exposed to high doses of neotame. These results confirm the need for further studies to use more reference genes, perform protein level measurements, and evaluate the mechanism of the relevant signaling pathways.

کلیدواژه‌ها [English]

  • dopamine
  • hepatotoxicity
  • male rats
  • neotame
  • qRT-PCR

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مجله بیوتکنولوژی کشاورزی
دوره 18، شماره 4
دی 1405
صفحه 629-648

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