INTRAMUSCULAR MESENCHYMAL STEM CELL APPLICATION WITH BORIC ACID REDUCE LDH DURING AN EXHAUSTIVE SWIM EXERCISE

Technical Summary: Intramuscular Mesenchymal Stem Cell Application with Boric Acid Reduces LDH During Exhaustive Swim Exercise

Problem Statement
Intense physical exercise induces skeletal muscle damage, characterized by the depletion of energy substrates, accumulation of metabolic byproducts (such as lactate), and the release of intracellular enzymes like Lactate Dehydrogenase (LDH) into the bloodstream. This damage leads to muscle fatigue, reduced athletic performance, and delayed recovery. While regenerative therapies using Mesenchymal Stem Cells (MSCs) show promise for tissue repair, challenges remain regarding cell viability, proliferation, and the efficacy of transplantation. Furthermore, while Boric Acid (BA) is known to influence hormone regulation and bone health, its specific role in enhancing MSC transplantation and mitigating exercise-induced muscle fatigue remains under-investigated. This study addresses the need for novel agents to enhance MSC effectiveness and improve rapid recovery strategies for athletes.

Methodology
The study utilized 30 female Wistar albino rats, divided into six groups (n=6 per group): Control, Exercise (Ex), Ex + Boric Acid (BA), Ex + MSCs, Ex + MSCs + BA (administered after exercise), and Ex + MSCs + BA (administered before exercise). An additional 5 rats were used for bone marrow MSC isolation.

• Exercise Protocol: Rats underwent an exhaustive swim exercise regimen over 15 days, divided into three periods. Period 1 was an adaptation phase; Periods 2 and 3 involved swimming until exhaustion. Exhaustion time was recorded daily.
• Intervention: Bone marrow-derived MSCs (1x10⁶ cells/1ml) were isolated and cultured. Boric acid doses were first optimized in cell culture to ensure cell viability. Intramuscular injections of MSCs and/or BA (10µg) were administered into the gastrocnemius muscles. The timing of administration varied by group (e.g., immediately after the second period of exercise).
• Measurements:
  • Physiological: Exhaustion time, body weight, and body temperature were monitored.
  • Biochemical: Plasma samples were collected 24 hours post-exercise to measure Lactate and Lactate Dehydrogenase (LDH) levels.
  • Histological: Gastrocnemius-soleus muscle tissues were fixed, sectioned, and stained with Hematoxylin & Eosin (H&E). Analysis focused on leukocyte infiltration, muscle fiber integrity, Z-band fluctuations, and the presence of centrally located nuclei (indicative of regeneration).
• Statistical Analysis: Data were analyzed using Student's t-test, Mann–Whitney U-test, and One-way ANOVA with post hoc corrections. Significance was set at p < 0.05.

Key Results

• Exhaustion Time: The combination of MSCs and BA significantly increased exhaustion time compared to the exercise-only group. Specifically, administering MSCs and BA before exercise increased exhaustion time by approximately 6-fold, while administration after exercise increased it by 4.5-fold. BA alone increased time by 2.5-fold, and MSCs alone by 4-fold.
• Lactate Dehydrogenase (LDH): The study found that the combination of MSCs and BA reduced LDH levels following exhaustive exercise, indicating a reduction in muscle membrane damage. However, the abstract notes a statistical nuance where the reduction in LDH for the combined group was reported as p>0.05 in the abstract text but described as a beneficial reduction in the discussion and conclusion.
• Lactate Levels: Contrary to the traditional view that high lactate indicates fatigue, the study observed that groups receiving MSCs and BA had significantly increased lactate levels (p=0.05). The authors interpret this as a sign that fatigue onset was delayed, allowing for prolonged exercise and higher lactate production before exhaustion.
• Histological Findings: The Exercise group showed significant muscle fiber disruption, leukocyte infiltration, and loss of integrity. In contrast, the MSCs and MSCs+BA groups exhibited muscle regeneration, evidenced by centrally located nuclei and muscle fibers that appeared closer to the normal structure of the control group.
• Physical Characteristics: Body temperature was significantly lower in groups receiving BA and MSCs compared to the exercise-only group, suggesting a mitigation of exercise-induced hyperthermia.

Significance and Claims
The authors claim this is the first study to investigate the intramuscular application of Boric Acid in combination with Mesenchymal Stem Cells for treating exercise-induced muscle damage. The primary significance of the work lies in demonstrating that:

1. Enhanced Endurance: The MSCs-BA combination significantly improves swimming endurance and delays the onset of fatigue.
2. Muscle Regeneration: Histological evidence confirms that this combination promotes muscle regeneration, as seen by the presence of centrally located nuclei and reduced tissue damage.
3. Protective Mechanism: The treatment appears to protect muscle tissue from the damage associated with exhaustive exercise, as indicated by lower LDH levels and reduced histological damage scores.
4. Novel Therapeutic Approach: The study suggests that Boric Acid can serve as a supportive agent to enhance the viability and therapeutic potential of MSCs in cellular therapy for sports recovery.

The paper concludes that while further studies with different doses and combinations are necessary, intramuscular administration of Boric Acid with MSCs is a beneficial strategy for improving athletic performance and accelerating recovery from exhaustive exercise.