The Effects of 5-methyltetrahydrofolate Supplementation in Patients With Metabolic Dysfunction-Associated Steatotic Liver Disease (MASLD)

July 30, 2026 updated by: Fatemeh Tamjid, Tabriz University of Medical Sciences

The Effect of 5-methyltetrahydrofolate Supplementation on Serum Folate and Homocysteine Level and PPARα and TNFα Gene Expression in Patients With Metabolic Dysfunction-Associated Steatotic Liver Disease: a Double-blind, Parallel Randomized Controlled Trial Study

To determine the effect of MTHF supplementation on serum folate and homocysteine level, metabolic, nutritional status, liver function, and PPARα and TNFα gene expression in patients with MASLD

Study Overview

Status

Not yet recruiting

Conditions

Intervention / Treatment

Detailed Description

Metabolic dysfunction-associated steatotic liver disease (MASLD), formerly termed non-alcoholic fatty liver disease (NAFLD), is diagnosed via liver biopsy or imaging when steatosis is present in the absence of alcohol intake or other hepatic disorders. As the liver manifestation of metabolic syndrome, it commonly coexists with obesity, diabetes, dyslipidemia, hypertension, and related conditions. Global prevalence of MASLD continues to rise.

Evidence from an earlier systematic review and meta-analysis indicated that MASLD patients had significantly lower serum folate and higher homocysteine concentrations. Folate is an essential water-soluble B vitamin that occurs in multiple chemically related forms. Food folates are mainly reduced and polyglutamated, with 5-MTHF predominating in both the diet and systemic circulation. 5-MTHF does not require reduction by DHFR and can enter the bloodstream directly for use. Reduced folates act as methyl donors in one carbon metabolism, supporting cellular proliferation, homocysteine re-methylation to methionine, nucleic acid synthesis and methylation of DNA, RNA, proteins and phospholipids.

Experimental studies have demonstrated that diet-induced hyperhomocysteinemia promotes hepatic steatosis and liver injury and folate as a key regulator of homocysteine concentration, may exert hepatoprotective effects. Evidence suggests that folate may improve hepatic lipid metabolism by activating peroxisome proliferator-activated receptor alpha (PPARα) signaling and modulate the immune response and reduce inflammatory mediators. Nevertheless, no evidence on the effects of folate on PPARα and TNFα gene expression in MASLD patients exist. Moreover, PPARα gene expression is dysregulated in MASLD and related metabolic conditions; PPARα is highly expressed in the liver, skeletal muscle and brown adipose tissue, stimulates β-oxidation and suppresses fatty-acid synthesis. Although the effect of 5-MTHF supplementation on gene expression of PPARα and TNFα in MASLD patients has not been examined, evidence showed that folate can modulate PPARα and TNFα. As folate has been shown to affect lipid metabolism and inflammation, we hypothesized that 5-MTHF supplementation might regulate PPARα and TNFα expression in MASLD patients. This randomized, double-blind, placebo-controlled clinical trial will therefore be undertaken to determine the effects of 5-MTHF supplementation on serum levels of folate and homocysteine, and gene expression of PPARα and TNFα in MASLD patients.

Study Type

Interventional

Enrollment (Estimated)

44

Phase

  • Not Applicable

Contacts and Locations

This section provides the contact details for those conducting the study, and information on where this study is being conducted.

Study Contact

Study Contact Backup

Study Locations

Participation Criteria

Researchers look for people who fit a certain description, called eligibility criteria. Some examples of these criteria are a person's general health condition or prior treatments.

Eligibility Criteria

Ages Eligible for Study

  • Adult

Accepts Healthy Volunteers

No

Description

Inclusion Criteria:

  • Adult men or women (18-50 years)
  • Diagnosis of MASLD (grade 1 or 2 of steatosis confirmed by ultrasound)
  • Body mass index (BMI) = 25-34.9 kg/m²
  • Providing written informed consent

Exclusion Criteria:

  • Pregnancy, lactation, or plans to get pregnant during the next three months.
  • Liver disease (viral hepatitis, autoimmune liver disease, cirrhosis, drug-induced hepatotoxicity, or alcoholic fatty liver disease), heart or renal failure, kidney stones, any neoplasia, inflammatory disease, hypothyroidism, hypercortisolism, or hypertension
  • Taking drugs affecting glucose or lipid metabolism, folate supplements, anti-obesity medications, weight-loss diets, or dietary supplements
  • Lifestyle factors known to impact folate status (current smoking, alcohol intake, recreational drug use)
  • Pre-existing conditions affecting folate status (malabsorptive or inflammatory bowel diseases, active celiac disease, gastric bypass surgery, atrophic gastritis, epilepsy, advanced liver disease, kidney dialysis, type 1 or 2 diabetes mellitus, or sickle cell trait/anemia)
  • Medications that interfere with B-vitamin metabolism (chloramphenicol, methotrexate, metformin, sulfasalazine, phenobarbital, phenytoin, primidone, triamterene, barbiturates)

Study Plan

This section provides details of the study plan, including how the study is designed and what the study is measuring.

How is the study designed?

Design Details

  • Primary Purpose: Treatment
  • Allocation: Randomized
  • Interventional Model: Parallel Assignment
  • Masking: Quadruple

Arms and Interventions

Participant Group / Arm
Intervention / Treatment
Placebo Comparator: Placebo
Dietary supplement: placebo group
Patients in this group will receive placebo for 90 days. The placebo is corn starch/ cellulose and will be consumed once a day. Placebo tablets will be manufactured by Ashbal Chemi Co. (Tehran, Iran).
Experimental: Intervention
Dietary supplement: 5-methyltetrahydrofolate (5-MTHF)
Patients in this group will receive 5-methyltetrahydrofolate tablets (800 mcg) once a day for 90 days. Tablets will be manufactured by Ashbal Chemi pharmaceutical company (Qfol, Ashbal Chemi Co., Tehran, Iran).

What is the study measuring?

Primary Outcome Measures

Outcome Measure
Measure Description
Time Frame
Serum folate level
Time Frame: 3 months
Changes in serum folate level pre and post the 3-month intervention period.
3 months
Serum homocysteine level
Time Frame: 3 months
Changes in serum homocysteine level pre and post the 3-month intervention period.
3 months
Expression of PPARα and TNFα genes
Time Frame: 3 months
Changes in expression of PPARα and TNFα genes pre and post the 3-month intervention period.
3 months

Secondary Outcome Measures

Outcome Measure
Measure Description
Time Frame
Liver biochemical parameters (ALT (alanine aminotransferase), AST (aspartate aminotransferase), and GGT (gamma-glutamyl transferase)
Time Frame: 3 months
Changes in ALT, AST, and GGT pre and post the 3-month intervention period.
3 months
The fibrosis-4 (FIB-4) index
Time Frame: 3 months
Changes in FIB-4 index pre and post the 3-month intervention period. The Fibrosis-4 (FIB-4) index will be calculated using the following formula: FIB-4 = (Age [years] × AST [U/L]) / (Platelet count [10⁹/L] × √ALT [U/L]). FIB-4 values >1.3, indicate a greater likelihood of liver fibrosis.
3 months
Quality of life using SF-36 (36-Item Short Form Health Survey) questionnaires
Time Frame: 3 months
Changes in quality-of-life pre and post the 3-month intervention period. Health-related quality of life will be assessed using the validated 36-Item Short Form Health Survey (SF-36). The questionnaire evaluates eight health domains: physical functioning, role limitations due to physical health, bodily pain, general health, vitality, social functioning, role limitations due to emotional problems, and mental health. Scores for each domain will be transformed to a 0-100 scale according to the standard scoring algorithm, with higher scores indicating better health-related quality of life.
3 months
Lipid profile (triglycerides, total cholesterol, LDL-C (low-density lipoprotein cholesterol), HDL-C (high-density lipoprotein cholesterol))
Time Frame: 3 monhs
Changes in lipid profile (triglycerides, total cholesterol, LDL-C, HDL-C) pre and post the 3-month intervention period.
3 monhs
Fasting blood glucose
Time Frame: 3 months
Changes in fasting blood glucose pre and post the 3-month intervention period.
3 months
Fasting serum insulin
Time Frame: 3 months
Changes in fasting serum insulin pre and post the 3-month intervention period.
3 months
QUICKI (quantitative insulin sensitivity check index)
Time Frame: 3 months
Changes in QUICKI pre and post the 3-month intervention period. The quantitative insulin sensitivity check index (QUICKI) will be calculated as 1/[log(fasting insulin [µU/mL]) + log(fasting glucose [mg/dL])], higher values indicating greater insulin sensitivity.
3 months
HOMA-IR (homeostatic model assessment of insulin resistance
Time Frame: 3 months
Changes in HOMA-IR pre and post the 3-month intervention period. Insulin resistance will be assessed using the homeostatic model assessment of insulin resistance (HOMA-IR), calculated as fasting insulin (µU/mL) × fasting glucose (mg/dL) / 405, higher values indicating greater insulin resistance.
3 months
Weight
Time Frame: 3 months
Changes in weight pre and post the 3-month intervention period.
3 months
Body Mass Index (BMI)
Time Frame: 3 months
Changes in BMI pre and post the 3-month intervention period. Body mass index (BMI) will be calculated as weight (kg) divided by the square of height (m²) and expressed as kg/m².
3 months
Waist circumference
Time Frame: 3 months
Changes in waist circumference pre and post the 3-month intervention period.
3 months
Waist-to-hip ratio (WHR)
Time Frame: 3 months
Changes in WHR pre and post the 3-month intervention period. Waist-to-hip ratio (WHR) will be calculated by dividing waist circumference by hip circumference.
3 months
Body composition (fat-free mass)
Time Frame: 3 months
Changes in fat-free mass (%) pre and post the 3-month intervention period. Body composition will be determined using a bioelectrical impedance analyzer.
3 months
Body composition (fat mass)
Time Frame: 3 months
Changes in fat mass (%) pre and post the 3-month intervention period. Body composition will be determined using a bioelectrical impedance analyzer.
3 months

Collaborators and Investigators

This is where you will find people and organizations involved with this study.

Sponsor

Tabriz University of Medical Sciences

Investigators

  • Principal Investigator: Bahram Pourghassem Gargari, Tabriz University of Medical Sciences

Study record dates

These dates track the progress of study record and summary results submissions to ClinicalTrials.gov. Study records and reported results are reviewed by the National Library of Medicine (NLM) to make sure they meet specific quality control standards before being posted on the public website.

Study Major Dates

Study Start (Estimated)

September 30, 2026

Primary Completion (Estimated)

August 30, 2027

Study Completion (Estimated)

August 30, 2027

Study Registration Dates

First Submitted

July 22, 2026

First Submitted That Met QC Criteria

July 30, 2026

First Posted (Actual)

July 31, 2026

Study Record Updates

Last Update Posted (Actual)

July 31, 2026

Last Update Submitted That Met QC Criteria

July 30, 2026

Last Verified

July 1, 2026

More Information

Terms related to this study

Keywords

Additional Relevant MeSH Terms

Other Study ID Numbers

  • IRCT20100123003140N26

Plan for Individual participant data (IPD)

Plan to Share Individual Participant Data (IPD)?

YES

Drug and device information, study documents

Studies a U.S. FDA-regulated drug product

No

Studies a U.S. FDA-regulated device product

No

This information was retrieved directly from the website clinicaltrials.gov without any changes. If you have any requests to change, remove or update your study details, please contact register@clinicaltrials.gov. As soon as a change is implemented on clinicaltrials.gov, this will be updated automatically on our website as well.

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