Identification of Functional Cellular Markers Related to Human Health, Frailty and Chronological Age

Abstract

Aging leads to a decline in physiological reserves, an increase in age-related diseases, reduced functional ability and a shortened healthspan. While molecular markers of chronological aging exist, their link to general health and intrinsic capacity (IC), a composite measure of physical and mental capacities, remains unclear. This study integrates the WHO's Healthy Aging framework with geroscience to explore fibroblasts as indicators of health. We assessed primary skin fibroblasts from 133 individuals aged 20-96, evaluating their ability to maintain tissue structure, modulate immune responses and regulate metabolism (SIM functions). By combining functional and molecular analyses, we investigated the relationship between fibroblast performance, chronological age and IC. Our results demonstrate that fibroblast SIM functions are modified with stressors and age, correlating with IC rather than just chronological age. Notably, fibroblasts from pre-frail and frail individuals exhibited reduced mitochondrial respiration and lower extracellular periostin levels, with periostin being able to capture IC status, irrespective of age and sex, reflecting a cellular 'health memory'. The SIM paradigm provides a complementary framework to the established hallmarks of aging, advancing our understanding of how cellular aging impacts functional decline. These findings suggest that fibroblast-derived markers could serve as indicators of frailty and reduced IC, enabling early detection of individuals at risk for health deterioration and laying the foundation for early identification of functional decline.

Keywords: cellular aging; dermal fibroblast; extracellular matrix; healthy aging; intrinsic capacity; metabolism.

FIGURE 1

Cellular characteristics of human skin fibroblasts that are significantly correlated with chronological age. Linear regression with marginal distribution represents cell parameters as a function of chronological age. Correlations between age and cell doubling time (days) (A), number of γH2AX foci per cell (B), extracellular IL‐6 concentration (pg/mL/10⁵ cells) (C) and number of p16 spots per cell (D) are shown. The association of senescence associated‐β‐galactosidase (MFI) (E) with age after doxorubicin challenge is shown. The black line represents the regression line, and the dashed line shows the 95% confidence of the fit. Histograms depict the marginal distribution of the respective variable. r and p‐value represent the Pearson correlation coefficient and the associated p‐value for each measured parameter with age. A p‐value < 0.05 was considered significant (A–E).

FIGURE 2

Overview of the study design using dermal fibroblasts from 133 donors selected from the INSPIRE T cohort (ages 20–96 years). Cell characteristics were examined on skin fibroblasts according to the three transverse and interconnected functions—tissue Structuration, Inflammation and the ability to regulate Metabolism (SIM) and senescence. The outcomes measured results per experiments are listed.

FIGURE 3

Association of homeostatic dysregulation with chronological age. Scatterplot of homeostatic dysregulation (MD) versus age for n = 108 donors with complete data on 31 cellular markers representing: (A) stroma/structure (% CFU‐F, extracellular Periostin, cell migration basal/residual chemoattractant), inflammation (extracellular IL1‐β basal/LPS residual, extracellular IL‐6 basal/LPS residual, extracellular IL‐10 basal/LPS residual, extracellular TGF‐β basal/LPS residual, extracellular IFN‐β basal/Poly I:C residual), metabolism (% differentiated adipocytes, differentiation intensity, OCR‐basal respiration, OCR‐uncoupled respiration and basal ECAR) and senescence (cell size basal/doxorubicin residual, beta galactosidase basal/doxorubicin residual, number of yH2AX spots per cell basal/doxorubicin residual, number of p16 spots per cell basal/doxorubicin residual, nucleus area basal/doxorubicin residual, cell granularity basal/doxorubicin residual) and (B) with SIM markers without senescence. The black line represents the regression line and the dashed lines show the 95% confidence of the fit. Histograms depict the marginal distribution of the respective variable. r and p‐value represent the Pearson correlation coefficient, and the associated p‐value for each measured parameter with age. A p‐value < 0.05 was considered significant.

FIGURE 4

The chronological age affects the regulation of pivotal molecular mediators implicated in the organization of stroma/structure, inflammation and metabolism within skin fibroblasts. Linear regression with marginal distribution represents cell parameters from stroma/structure (A, B), inflammation (C–E) and metabolism (F, G) as a function of chronological age. Correlation between age and TIMP1 mRNA expression (2^−ΔCt) (A) and extracellular Periostin logarithmic concentration (pg/mL/10⁵ cells) (B) are shown. Association with age of extracellular IL‐6 (C) and IL1‐β (D) concentration (pg/mL/10⁵ cells) after LPS stimulation (1 μg/mL) and extracellular IFN‐β concentration (pg/mL/10⁵ cells) after Poly I:C stimulation (100 μg/mL) (E) are shown. Correlation between age and uncoupled respiration over basal mitochondrial respiration (%) (F), % differentiated cells (adipocytes) (G) HEXOKINASE 2 mRNA expression (2^−ΔCt) (H), mRNA expression (2^−ΔCt) of SOD1 (I) and GPX1 (J) are shown. The black line represents the regression line, and the dashed line shows the 95% confidence of the fit. Histograms depict the marginal distribution of the respective variable. r and p‐value represent the Pearson correlation coefficient and the associated p‐value for each measured parameter with age. A p‐value < 0.05 was considered significant (A–J).

FIGURE 5

Metabolism and structure human skin fibroblast markers reveal functional decline regardless of chronological age. Violin plots show the distribution of basal and maximal mitochondrial respiration (pmol/min/2.10⁴ cells) (A, B), CD36 mRNA expression (C), % CFU‐f (D) and extracellular Periostin concentration (pg/mL/10⁵ cells) (E) in robust and pre‐frail/frail population. The horizontal line represents the median. Logistic regressions were performed to examine the association between cell parameters and pre‐frailty/frailty status. Cell parameters p values ‘p _adj’ are based on logistic regression with adjustment for age and sex (A–E). Linear regression with marginal distribution represents Intrinsic Capacity score (IC) as a function of extracellular Periostin concentration (pg/mL/10⁵cells) and extracellular Periostin p value ‘p _adj’ is based on linear regression with adjustment for age and sex. The black line represents the regression line and the dashed line show the 95% confidence of the fit. Histograms depict the marginal distribution of the respective variable (F). Violin plot shows the distribution of extracellular PeriostinN concentration (pg/mL/10⁵ cells) by IC centile group. The horizontal line represents the median. Association between IC centile groups (age and sex specific) and extracellular PERIOSTIN secretion is determined using one‐way ANOVA test (G). A p‐value < 0.05 was considered significant (A–G).