12 Weeks of Combined Endurance and Resistance Training Reduces Innate Markers of Inflammation in a Randomized Controlled Clinical Trial in Patients with Multiple Sclerosis

The researchers tested a supervised, combined exercise program consisting of both endurance (aerobic) training and resistance (strength) training, performed three times per week for 12 weeks. The comparator was a control group that continued their usual daily activities and received no exercise intervention. Both groups continued their standard medical treatments for Multiple Sclerosis (MS) throughout the study.

The primary outcomes were changes in blood markers of innate immune system inflammation: tumour necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), interleukin-17 (IL-17), and C-reactive protein (CRP). Secondary outcomes included measures of physical fitness (VO₂max, muscle strength), body composition (body fat percentage, lean mass), and self-reported quality of life.

The exercise sessions lasted approximately 60–70 minutes each. The endurance component involved cycling on a stationary bike at 60–80% of heart rate reserve (a moderate-to-vigorous intensity), starting at 20 minutes per session and progressing to 40 minutes by week 12. The resistance component involved 8–10 exercises targeting major muscle groups (leg press, chest press, lat pulldown, shoulder press, leg extension, leg curl, bicep curl, tricep extension, and abdominal exercises), performed at 60–80% of one-repetition maximum (1RM), for 2–3 sets of 8–12 repetitions per exercise.

The study enrolled 34 adults diagnosed with relapsing-remitting Multiple Sclerosis (RRMS), the most common form of MS characterised by periods of symptom flare-ups followed by partial or full recovery. Participants were recruited from a university hospital neurology clinic in Iran.

Key inclusion criteria:

Confirmed diagnosis of RRMS according to McDonald criteria

Expanded Disability Status Scale (EDSS) score between 1.0 and 4.0 (mild to moderate disability — able to walk without aid or with minimal assistance)

No relapse or corticosteroid treatment in the 30 days before the study

No regular exercise training (defined as structured exercise more than once per week) in the previous 6 months

Age range: 20–50 years

Key exclusion criteria:

Cardiovascular, pulmonary, or orthopaedic conditions that would contraindicate exercise

Pregnancy

Cognitive impairment that would prevent understanding of instructions

Use of immunosuppressive medications other than standard disease-modifying therapies (interferon beta or glatiramer acetate)

The final sample consisted of 34 participants (17 per group), with a mean age of approximately 34 years (range 22–48). The majority were female (roughly 70%, consistent with the higher prevalence of MS in women). Average disease duration was about 5–6 years, and average EDSS score was approximately 2.5 (mild disability — able to walk independently but with some limitations).

Blood markers of inflammation were measured from venous blood samples collected after an overnight fast (at least 10 hours) and at least 48 hours after the last exercise session to avoid acute exercise-induced inflammation. The following assays were used:

**TNF-α and IL-6:** Measured using enzyme-linked immunosorbent assay (ELISA) kits with sensitivity reported as 0.5 pg/mL and 0.7 pg/mL respectively. These are standard, commercially available kits with good reliability.

**IL-17:** Measured using a high-sensitivity ELISA kit (sensitivity 0.5 pg/mL). IL-17 is a pro-inflammatory cytokine particularly relevant to autoimmune diseases like MS.

**CRP:** Measured using a high-sensitivity immunoturbidimetric assay (sensitivity 0.1 mg/L). High-sensitivity CRP is the standard clinical test for low-grade systemic inflammation.

Physical fitness was assessed using:

**VO₂max:** Estimated from a submaximal cycle ergometer test using the Astrand-Ryhming protocol, which predicts maximal oxygen uptake from heart rate response at a submaximal workload. This is a validated field test but less accurate than direct gas exchange measurement.

**Muscle strength:** Measured as 1-repetition maximum (1RM) for leg press and chest press, using standard resistance training equipment. This is the gold standard for assessing maximal strength.

**Body composition:** Measured using bioelectrical impedance analysis (BIA), which estimates body fat percentage and lean mass by passing a small electrical current through the body. BIA is convenient but less accurate than DEXA scanning.

Quality of life was assessed using the Multiple Sclerosis Quality of Life-54 (MSQOL-54) questionnaire, a validated instrument specific to MS that covers physical and mental health domains.

**Study design:** This was a randomised controlled clinical trial (RCT) with two parallel groups: exercise intervention and usual-care control. Participants were randomly assigned to either group using a computer-generated random number sequence.

**Randomisation:** The randomisation sequence was generated by a statistician not involved in the study. Allocation concealment was achieved using sequentially numbered, opaque, sealed envelopes — meaning the person enrolling participants could not predict which group the next participant would be assigned to. This is a strong method for preventing selection bias.

**Blinding:** This study was not blinded. Participants knew whether they were exercising or not (impossible to blind in an exercise intervention), and the outcome assessors (the researchers measuring blood markers and fitness tests) were not blinded either. This is a significant methodological weakness because expectation effects and observer bias could influence results, particularly for subjective outcomes like quality of life. However, the primary outcomes (blood markers) are objective laboratory measures, which are less susceptible to bias than self-reported outcomes.

**Duration:** The intervention lasted 12 weeks, with exercise sessions performed three times per week (36 total sessions). Assessments were conducted at baseline (week 0) and post-intervention (week 12). There was no follow-up period after the intervention ended, so the durability of any effects is unknown.

**Adherence and supervision:** All exercise sessions were supervised by an exercise physiologist or physiotherapist. Adherence was recorded, and participants who missed more than two consecutive sessions or more than 20% of total sessions were excluded from the final analysis. The authors reported that 17 out of 20 participants in the exercise group completed the study (85% adherence), and 17 out of 19 in the control group completed (89% retention). Three participants dropped out of the exercise group (two due to personal reasons, one due to an MS relapse unrelated to exercise) and two dropped out of the control group (one due to relocation, one due to personal reasons).

**Statistical approach:** The researchers used analysis of covariance (ANCOVA) to compare post-intervention values between groups, adjusting for baseline values. This is appropriate for RCTs because it accounts for any baseline differences between groups (even after randomisation, some differences can occur by chance). Effect sizes were reported as partial eta-squared (η²p), which indicates the proportion of variance explained by the intervention. They also reported within-group changes using paired t-tests. A p-value of less than 0.05 was considered statistically significant.

**What this design can and cannot prove:**

This RCT design can establish causality — because participants were randomly assigned, any differences between groups at the end of the study can be attributed to the exercise intervention rather than to pre-existing differences between participants. This is the strongest design for testing whether exercise *causes* changes in inflammation.

However, the lack of blinding means that some outcomes (particularly quality of life) could be influenced by placebo effects — participants who know they are exercising may report feeling better simply because they expect to. The objective blood markers are less vulnerable to this, but not entirely immune (stress from expectation could theoretically affect some inflammatory markers).

The design cannot tell us:

Whether the effects persist after stopping exercise (no follow-up)

Whether the effects are due to the endurance component, the resistance component, or their combination (no separate groups for each type of exercise)

Whether the effects generalise to people with more severe MS (EDSS > 4.0) or other forms of MS (progressive MS)

The mechanisms by which exercise reduces inflammation (the study measured markers but did not investigate pathways like changes in adipose tissue, stress hormones, or immune cell trafficking)

**Major methodological weaknesses:**

1. Small sample size (n=34 total, 17 per group) — increases risk of Type II error (failing to detect real effects) and makes results less reliable

2. No blinding of participants or assessors

3. No sham exercise control (e.g., stretching or light activity)

4. No intention-to-treat analysis (they only analysed completers, which can bias results if dropouts differ from completers)

5. Single-centre study in Iran — may not generalise to other populations or healthcare systems

6. No adjustment for multiple comparisons (testing several inflammatory markers increases the chance of false positives)

**Primary outcomes (inflammatory markers):**

**TNF-α:** Decreased by 28.6% in the exercise group (from 18.2 ± 4.1 pg/mL to 13.0 ± 3.8 pg/mL) compared to a 3.8% increase in the control group (from 17.9 ± 4.3 pg/mL to 18.6 ± 4.5 pg/mL). The between-group difference was statistically significant (p = 0.003, η²p = 0.28 — a large effect size, meaning 28% of the variance in TNF-α change was explained by group assignment).

**IL-6:** Decreased by 22.7% in the exercise group (from 4.4 ± 1.2 pg/mL to 3.4 ± 1.0 pg/mL) compared to a 2.3% increase in the control group (from 4.3 ± 1.1 pg/mL to 4.4 ± 1.2 pg/mL). The between-group difference was statistically significant (p = 0.008, η²p = 0.22 — large effect size).

**IL-17:** Decreased by 19.4% in the exercise group (from 6.7 ± 1.8 pg/mL to 5.4 ± 1.5 pg/mL) compared to a 1.5% increase in the control group (from 6.6 ± 1.7 pg/mL to 6.7 ± 1.8 pg/mL). The between-group difference was statistically significant (p = 0.012, η²p = 0.19 — large effect size).

**CRP:** Decreased by 38.5% in the exercise group (from 2.6 ± 0.9 mg/L to 1.6 ± 0.7 mg/L) compared to a 3.8% increase in the control group (from 2.5 ± 0.8 mg/L to 2.6 ± 0.9 mg/L). The between-group difference was statistically significant (p = 0.001, η²p = 0.32 — very large effect size).

**Secondary outcomes:**

**VO₂max:** Increased by 15.2% in the exercise group (from 28.9 ± 4.2 mL/kg/min to 33.3 ± 4.5 mL/kg/min) compared to no significant change in the control group (from 29.1 ± 4.0 mL/kg/min to 28.8 ± 4.1 mL/kg/min). Between-group difference significant (p < 0.001).

**Leg press 1RM:** Increased by 32.1% in the exercise group (from 68.4 ± 12.3 kg to 90.4 ± 14.1 kg) compared to no significant change in the control group (from 67.9 ± 11.8 kg to 68.5 ± 12.0 kg). Between-group difference significant (p < 0.001).

**Chest press 1RM:** Increased by 28.6% in the exercise group (from 35.7 ± 8.2 kg to 45.9 ± 9.5 kg) compared to no significant change in the control group (from 36.1 ± 7.9 kg to 36.4 ± 8.1 kg). Between-group difference significant (p < 0.001).

**Body fat percentage:** Decreased by 6.8% in the exercise group (from 32.4 ± 5.1% to 30.2 ± 4.8%) compared to no significant change in the control group (from 32.1 ± 4.9% to 32.3 ± 5.0%). Between-group difference significant (p = 0.015).

**Lean mass:** Increased by 3.1% in the exercise group (from 44.8 ± 6.2 kg to 46.2 ± 6.4 kg) compared to no significant change in the control group (from 45.1 ± 6.0 kg to 44.9 ± 6.1 kg). Between-group difference significant (p = 0.021).

**Quality of life (MSQOL-54):** The physical health composite score improved by 14.2% in the exercise group (from 58.3 ± 12.1 to 66.6 ± 11.8) compared to no significant change in the control group (from 57.9 ± 11.6 to 58.4 ± 12.0). The mental health composite score improved by 11.7% in the exercise group (from 61.5 ± 13.4 to 68.7 ± 12.9) compared to no significant change in the control group (from 62.1 ± 12.8 to 61.8 ± 13.1). Both between-group differences were significant (p < 0.01).

To translate these numbers into plain English:

**CRP reduction (~39%):** This is a substantial drop. For context, a CRP level of 2.6 mg/L (the baseline in this study) is considered moderate cardiovascular risk; dropping to 1.6 mg/L moves into the low-risk category. This is roughly equivalent to the reduction seen with statin therapy or significant weight loss (10% of body weight).

**TNF-α reduction (~29%):** This is a large effect for a cytokine that is notoriously difficult to modulate with lifestyle interventions. For comparison, anti-TNF biologic drugs (like adalimumab or etanercept) typically reduce TNF-α by 50–80%, but these are powerful pharmaceuticals with significant side effects. A 29% reduction from exercise alone is clinically meaningful.

**IL-6 reduction (~23%):** This is comparable to the reduction seen with regular moderate exercise in healthy populations (typically 10–25% in meta-analyses). IL-6 is a pleiotropic cytokine — it can be both pro- and anti-inflammatory depending on context — so the direction of change matters. In chronic inflammatory conditions, lower IL-6 is generally beneficial.

**VO₂max increase (~15%):** This is a clinically meaningful improvement. For a 34-year-old with MS starting at 28.9 mL/kg/min (which is "poor" cardiorespiratory fitness for that age), increasing to 33.3 mL/kg/min moves into the "fair" category. Each 1 MET (3.5 mL/kg/min) increase in cardiorespiratory fitness is associated with roughly 10–15% reduction in all-cause mortality in the general population.

**Strength increases (28–32%):** These are large improvements typical of a well-designed resistance training program in previously untrained individuals. For someone with MS, improved leg strength can translate directly to better walking ability, stair climbing, and fall prevention.