Reduced liver damage and fibrosis with combined SCD Probiotics and intermittent fasting in aged rat

Abstract

This study aimed to examine the impact of SCD Probiotics supplementation on liver biomolecule content and histological changes during a 30-day intermittent fasting (IF) program in 24-month-old male Sprague-Dawley rats. Rats underwent 18-h daily fasting and received 1 × 10⁸ CFU of SCD Probiotics daily. Liver tissue biomolecules were analysed using FTIR Spectroscopy, LDA, and SVM techniques, while histopathological evaluations used Haematoxylin and eosin and Masson trichrome-stained tissues. Blood samples were collected for biochemical analysis. Gross alterations in the quantity of biomolecules were observed with individual or combined treatments. LDA and SVM analyses demonstrated a high accuracy in differentiating control and treated groups. The combination treatments led to the most significant reduction in cholesterol ester (1740 cm^-1 ) and improved protein phosphorylation (A₁₂₃₉ /A₂₉₅₅ and A₁₀₈₀ /A₁₅₄₅ ) and carbonylation (A₁₇₄₀ /A₁₅₄₅ ). Individually, IF and SCD Probiotics were more effective in enhancing membrane dynamics (Bw₂₉₂₂ /Bw₂₉₅₅ ). In treated groups, histological evaluations showed decreased hepatocyte degeneration, lymphocyticinfiltration, steatosis and fibrosis. Serum ALP, LDH and albumin levels significantly increased in the SCD Probiotics and combined treatment groups. This study offers valuable insights into the potential mechanisms behind the beneficial effects of IF and SCD Probiotics on liver biomolecule content, contributing to the development of personalized nutrition and health strategies.

Keywords: FTIR; SCD probiotics; fibrosis; intermittent fasting; liver; rat.

FIGURE 1

Effects of intermittent fasting, SCD probiotics supplementation, and combined treatment on: (A) Body weight: Significant difference among groups (p < 0.0001). Comparisons: C versus F (p < 0.0001), C versus P (p = 0.1732), C versus FP (p = 0.0023), F versus P (p < 0.0001), F versus FP (p = 0.0418), and P versus FP (p = 0.0394), (B) Food consumption: Significant difference among groups (p < 0.0001). Comparisons: C versus F (p < 0.0001), C versus P (p = 0.1732), C versus FP (p < 0.0001), F versus P (p < 0.0001), F versus FP (p = 0.00538), and P versus FP (p < 0.0001), (C) Water consumption: No significant difference among groups (p = 0.607). C (control), F (Intermittent fasting), P (SDC Probiotics), and the FP applications (in which the Intermittent fasting and SCD Probiotics were applied together).

FIGURE 2

LDA discrimination plots for liver samples in the (A) full (4000–650 cm⁻¹) and (B) lipid (3000–2700 cm⁻¹) spectral regions. CLI (control), FLI (Intermittent Fasting), PLI (SDC Probiotics), and the FPLI applications (in which the intermittent fasting and SCD Probiotics were applied together).

FIGURE 3

Baseline‐corrected average infrared spectra in the 4000–650 cm⁻¹ spectral region for liver tissues. CLI (control), FLI (Intermittent Fasting), PLI (SDC Probiotics), and the FPLI applications (in which the intermittent fasting and SCD Probiotics were applied together).

FIGURE 4

The quantitative changes in liver‐associated spectrochemical parameters. The band areas for (A) lipid carbonyl (C=O stretchings, 1740 cm⁻¹), (B) Amide II (1545 cm⁻¹), (C) PO₂ antisymmetric (1239 cm⁻¹), and (D) PO₂ symmetric (1080 cm⁻¹). CLI (control), FLI (Intermittent Fasting), PLI (SDC Probiotics), and the FPLI applications (in which the intermittent fasting and SCD Probiotics were applied together).

FIGURE 5

The quantitative changes in liver‐associated spectrochemical parameters. The band area ratios for (A) acyl chain length of fatty acids (A₂₉₂₂/A₂₉₅₅), (B) protein phosphorylation (A₁₂₃₉/A₂₉₅₅), (C) protein phosphorylation (A₁₀₈₀/A₁₅₄₅), and (D) protein carbonylation (A₁₇₄₀/A₁₅₄₅). CLI (control), FLI (Intermittent Fasting), PLI (SDC Probiotics), and the FPLI applications (in which the intermittent fasting and SCD Probiotics were applied together).

FIGURE 6

Aged rat livers show increased steatosis and lymphocytic infiltration, but IF, SCD probiotics, and combination treatment groups improved these histopathological alterations (A). Representative images of haematoxylin and eosin staining of lymphocytic infiltration and bile duct proliferation areas. Black arrows show lymphocytic infiltration and green arrows show bile ducts (B). Black arrow heads show hepatic microvesicular steatosis. Representative graphics of quantification of lymphocytic infiltration area fraction (%) and hepatic microvesicular steatosis area fraction (%) in all groups (C). The squared areas in the respective microphotographs of all groups are enlarged in the upper right corner. Values are expressed as mean ± SEM; n = 7 rats in each group. p < 0.05, and p ≤ 0.0001*** (nonparametric Mann–Whitney U test). Scale bar = 100 μm. CLI (control), FLI (Intermittent Fasting), PLI (SDC Probiotics), and the FPLI applications (in which the intermittent fasting and SCD Probiotics were applied together).

FIGURE 7

Representative images from MT staining depict collagen deposition in the liver tissues of aged control rats, as well as those treated with IF, SCD probiotics, and a combination of both treatments. The quantification of the collagen density area fraction (%) for all groups is also provided (A). Close‐up images of specific areas highlighted by black arrows in the microphotographs from the previous figure are displayed (B). Values are expressed as mean ± SEM; n = 7 rats in each group. p ≤ 0.01*, and p ≤ 0.0001*** (nonparametric Mann–Whitney U test) versus control. Scale bar = 50 and 100 μm. CLI (control), FLI (Intermittent Fasting), PLI (SDC Probiotics), and the FPLI applications (in which the intermittent fasting and SCD Probiotics were applied together).