Conceptual framework and the study design.

We collected published datasets of human blood samples for gene expression, DNA methylation, and single-cell transcriptomic data. The analysis aimed to study the relation between the expression and DNA methylation of RTEs versus chronological and biological aging in large human cohorts. The single-cell transcriptomic datasets were employed for cell type-specific analysis of RTEs in PBMC to identify the relation between RTEs expression and ageing events for annotated cell types within old versus young PBMC samples.

Correlation analysis between RTEs expression, chronological age, and age-associated gene signature scores. a,

No correlation between RTEs expression and chronological age versus positive correlations between age-associated gene signature scores and LINE and LTR expressions. Pair-wise correlation coefficients were calculated between the expression of different RTE classes (LTR, LINE & SINE) and chronological age and six age-associated gene signature scores in monocytes (MESA), PBMCs (GARP), and the whole blood (GTP). b, Scatter plots displaying a positive correlation between LINE and LTR expressions and inflammaging, SASP, and senescence gene signature scores in PBMCs. c, Different families of RTEs were divided into two major groups based on their correlation and inverse correlation with age-associated signature scores in PBMC samples. d, Correlation matrix depicting all pair-wise combinations to identify the correlation between chronological age, RTE family expressions, and six age-associated signature scores in PBMCs. ** P ≤ 0.01, *** P ≤ 0.001, Pearson’s correlation. MESA, n=1202; GARP, n=139, GTP, n=359.

Upregulation of DNA repair vs inflammatory responses for samples with high expression of SINE vs LTR and LINE in the MESA and GARP cohorts.

a, GSVA demonstrates increased activity of DNA repair pathways in the group of samples with high vs low SINE expression in the MESA and GARP cohorts. In contrast, the inflammatory response is upregulated in the sample groups highly expressing LINE and LTR classes and families in the MESA and GARP cohorts. The samples in each cohort were divided into low (1st quartile), medium (2nd and 3rd quartile), and high (4th quartile) expression groups based on the expression of RTE classes or families. GSVA was applied on high vs low groups for each class and family of RTEs. The threshold for differential expression is set at |logFC| > 0.1 and P < 0.05 (online methods). b, The Radar plot shows the difference between the number of upregulated versus downregulated gene sets related to DNA repair and inflammatory response in each cohort. While high expression of SINE and Alu is significantly associated with high number of up-regulated DNA-repair gene sets, LINE and L1 expressions are highly related to high number of activated gene sets related to inflammatory response in the MESA and GARP cohorts. This result is not highly supported by the GTP cohort, more likely due to the low number of probes in this cohort.

Inverse correlation of DNA methylation levels of RTEs with the chronological age and the RTEs expression except for SINE expression.

a, b, Methylation levels of RTE classes inversely correlate with chronological age in monocyte (MESA) and whole blood (BSGS, SATSA, and GMPWAR) samples. Satellite DNA is included as a control group. c, Methylation levels versus low (1st quartile), medium (2nd and 3rd quartile), and high (4th quartile) expressions of RTE classes in monocytes (MESA). Wilcoxon test; ns: not significant. d, Correlation matrix for RTE expressions and methylation levels, and chronological age. * P ≤ 0.05, **** P ≤ 0.0001, Pearson’s correlation. MESA, n=1202; BSGS, n=614; GMPWAR, n=656; SATSA, n=1072.

Cell type-specific analysis of RTEs expression vs age-associated gene signature scores in two PBMC scRNA-seq cohorts.

a-f, Unique increased expression of RTEs in the Plasma B cells of the samples with high age-associated gene signature scores indicates the potential role of plasma B cells in ageing. Wilcoxon test. n = 21. g, Decreased expressions of RTE classes and increased age-associated gene signature scores in multiple annotated cell types obtained from the PBMCs of supercentenarians compared to ordinary elderlies as control. Supercentenarians, n=7; control, n=5, age 50s to 80s. Wilcoxon test was applied to identify the significant changes. NK, Natural killer cell; BC, B-cell; TC1, T-cell 1; TC2, T-cell 2; M14, CD14+ monocyte; M16, CD16+ monocyte; EC, Erythrocytes; MKI, MKI67+ proliferating cell; DC, Dendritic cell; MGK, Megakaryocyte.

Number of RTE-covering probes.

a, The number of microarray probes covering RTE regions in MESA, GTP, and GARP compared to the total number of RTE probes available in Illumina HumanHT-12 V4. b, The number of microarray probes covering RTE regions in MESA, GMPWAR, SATSA, BSGS compared to the total number of RTE probes available in Illumina Infinium 450k array.

Correlation analysis between age-associated gene signatures and RTE family expressions in human cohorts.

a, Weak correlation between a few RTE families and chronological age and strong positive correlation between L1, ERVL, ERVK, and MaLR with age-associated signature scores in PBMC samples (GARP cohort). b, c, Correlation matrix depicting all pair-wise combinations to identify the correlation between chronological age, RTE family expressions, and six age-associated gene expressions in monocytes (MESA) and the whole blood (GTP). * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001, Pearson’s correlation.

Comparing SINE expression in 25 cell types obtained from young versus old PBMC human samples.

No significant difference was observed in any cell type. The young group comprises healthy male donors aged 25 to 29, n=11; the old groups are healthy male donors aged 62 to 70, n=10; Wilcoxon test; ns: insignificant.

Comparing LINE expression in 25 cell types obtained from young versus old PBMC human samples.

No significant difference was observed in any cell type. The young group comprises healthy male donors aged 25 to 29, n=11; the old groups are healthy male donors aged 62 to 70, n=10—Wilcoxon test; ns: insignificant.

Comparing LTR expressions in 25 cell types obtained from young versus old PBMC human samples.

No significant difference was observed in any cell type. The young group comprises healthy male donors aged 25 to 29, n=11; the old groups are healthy male donors aged 62 to 70, n=10—Wilcoxon test; ns: insignificant.

Increasing trend of age-associated gene signature scores in high vs low LTR and LINE expression groups in the three human cohorts.

a,b The samples in each cohort were divided into (low (1st quartile), medium (2nd and 3rd quartile), and high (4th quartile) LTR and LINE expression groups, respectively. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001,

Different patterns of age-associated signature scores in MESA vs GARP vs GTP for high vs low SINE and Alu expression groups in the three human cohorts.

a, b, The samples in each cohort were divided into low (1st quartile), medium (2nd and 3rd quartile), and high (4th quartile) SINE and Alu expression groups, respectively. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤

Correlation of methylation levels of RTE families with chronological age in monocyte (MESA) and whole blood (BSGS, SATSA, and GMPWAR) samples.

a, b, Methylation levels of LTR and LINE/SINE families negatively correlate with chronological age in monocytes (MESA) and the whole blood (BSGS, SATSA, and GMPWAR). *** P ≤ 0.001, Wilcoxon test.

Methylation levels versus low (1st quartile), medium (2nd and 3rd quartile), and high (4th quartile) expressions of RTE families in monocytes.

While LINE families, MIR, and LTR families except ERVK show lower levels of methylation in higher expression groups, this pattern is not seen in Alu, CR1, and ERVl. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001, ns: not significant, Wilcoxon test.

Cell type-specific SINE expression versus low and high SASP score groups among 25 cell types in PBMC.

Plasma B cells demonstrate significantly elevated SINE expression in samples with high SASP scores. * P ≤ 0.05, ns: not significant, Pearson’s correlation.

Cell type-specific LINE expression versus low and high SASP score groups among 25 cell types in PBMC.

Plasma B cells demonstrate significantly elevated LINE expression in samples with high SASP scores. * P ≤ 0.05, ns: not significant, Pearson’s correlation.

Cell type-specific LTR expression versus low and high SASP score groups among 25 cell types in PBMC.

Plasma B cells demonstrate significantly elevated LTR expression in samples with high SASP scores. * P ≤ 0.05, ns: not significant, Pearson’s correlation.

Comparison of cell type-specific RTEs expression in supercentenarians versus normal aged cases.

* P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001, ns: not significant, Wilcoxon test.

Comparison of cell type-specific Senescence, inflammaging, and SASP gene signature scores in supercentenarians versus normal aged cases.

* P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001, ns: not significant, Wilcoxon test.

Comparison of cell type-specific Inflammatory cytokines, Inflammatory chemokines, and IFN-I gene signature scores in supercentenarians versus normal aged cases.

* P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001, ns: not significant, Wilcoxon test.