Mitigating Mitochondrial RNA Release During Aging to Control Inflammation and Senescence

Official title

Mitigating Mitochondrial RNA Release During Aging to Control Inflammation and Senescence, Preserving Organ Function and Enhancing Healthspan

Overview

The MIRACLE study aims to investigate age-related mitochondrial dysfunction, mitochondrial RNA (mtRNA) release, inflammation, and cellular senescence in adult participants across three age groups. Skin-derived fibroblasts and peripheral blood mononuclear cells (PBMCs) will be isolated from skin biopsy and blood samples to characterize age-related cellular and molecular changes and to test experimental therapeutic strategies identified in preclinical studies. Serum, plasma, and whole-blood RNA will be used for protocol-defined analyses of circulating inflammatory mediators and systemic transcriptional signatures related to inflammation, type I interferon activation, mitochondrial stress response, immune aging, and senescence-associated pathways.

Detailed description

Mitochondria are crucial for ATP production and intracellular signaling in higher eukaryotic cells. These organelles contain genetic material that reflects their bacterial ancestry, including mitochondrial RNA (mtRNA). Under normal conditions, mtRNA is tightly confined and processed within mitochondria, ensuring the proper synthesis of proteins required for oxidative phosphorylation. Recent evidence suggests that mitochondrial stress may promote the leakage of mitochondrial components, including mtRNA. Once released into the cytosol, mtRNA can be sensed by cytosolic pattern recognition receptors (PRRs), thereby activating signaling pathways that promote the production of pro-inflammatory cytokines.

Although these mechanisms have been investigated mainly under conditions of acute stress, the specific contribution of mitochondrial dysfunction and mtRNA release to chronic low-grade inflammation and cellular senescence during physiological aging remains incompletely understood. This represents an important knowledge gap in the understanding of molecular processes that may contribute to age-related inflammation and functional decline.

The MIRACLE project includes a broade series of preclinical in vitro and in vivo experiments aimed at clarifying the mechanisms linking mitochondrial dysfunction, mtRNA release, inflammatory pathway activation, and cellular senescence during aging. These experimental activities are designed to define the biological pathways involved and to identify potential strategies capable of modulating mtRNA-associated inflammatory and senescence responses.

Within this broader framework, the present human study is intended to corroborate and extend the preclinical findings in humans. To this aim, adult participants across different age groups will be enrolled, and skin biopsy and blood samples will be collected to obtain skin-derived fibroblasts, peripheral blood mononuclear cells (PBMCs), serum, plasma, and whole-blood RNA.

Skin-derived fibroblasts will provide an accessible primary cell model for the investigation of age-related cellular and molecular changes. Fibroblasts isolated from participants of different ages will be used to assess mitochondrial function, mtRNA release, inflammatory signaling, and markers of cellular senescence. PBMCs collected from the same participants will be analyzed as a complementary blood-derived cellular model to evaluate systemic immune and inflammatory features, including immunosenescence-related signatures. Serum and plasma samples will be used to measure circulating inflammatory mediators and senescence-associated factors, while whole-blood RNA will be used to assess systemic transcriptional signatures related to inflammation, type I interferon activation, mitochondrial stress response, immune aging, and senescence-associated pathways.

Interventions

• Procedure Skin biopsy and venous blood sampling
  A single 3-4 mm punch biopsy of forearm skin is performed under local anesthesia (lidocaine 1% with epinephrine 1:100,000) for primary dermal fibroblast isolation. Venous blood is collected by standard phlebotomy from the antecubital fossa (43 mL total volume per participant): 5 mL in SST tube for serum separation, 3 mL in EDTA tube for plasma separation, 32 mL in EDTA tubes for PBMC isolation by density gradient centrifugation, and 3 mL in Tempus Blood RNA tube for whole-blood RNA stabilization

Primary outcome measures

• Age-dependent differences in cytosolic release of mitochondrial RNA (mtRNA) in primary human fibroblasts [Time frame: Day 1 (at enrollment)]
• Age-dependent differences in mitochondrial function in primary human fibroblasts [Time frame: Day 1 (at enrollment)]
• Activation of inflammatory pathways in primary human fibroblasts [Time frame: Day 1 (at enrollment)]
• Activation of senescence pathways in primary human fibroblasts [Time frame: Day 1 (at enrollment)]
• Comparison of fibroblast findings with PBMC-derived parameters from the same participants [Time frame: Day 1 (at enrollment)]

Eligibility criteria

Inclusion criteria

• Male and female
• Age between 18 and 90 years (stratified in three groups: young, middle-aged, elderly)
• Written informed consent

Exclusion criteria

• Inability to understand the potential risk and benefits of the study
• Legal incapacity
• Subjects who have taken antibiotics, anti-inflammatory drugs, or antihistamines within the past 7 days
• Diagnosis of diabetes mellitus
• Use of anticoagulant medications
• Any subject with a contraindication to the mini-invasive biopsy procedure

Criteria are shown verbatim from the registry (in English). Final eligibility is always assessed by the study center.

Healthy volunteers: Yes

Study design

Allocation

Non-randomized

Model

Parallel assignment

Masking

Open label

Primary purpose

Basic science

Study locations

Italy · 2 centers

• Laboratory for Targeted Therapy in Autoimmune Diseases — Bergamo
• Centro di Ricerche Cliniche per le Malattie Rare "Aldo e Cele Daccò" — Ranica

Publications

• Zuber TJ. Punch biopsy of the skin. Am Fam Physician. 2002 Mar 15;65(6):1155-8, 1161-2, 1164. PMID 11925094
• Nischal U, Nischal Kc, Khopkar U. Techniques of skin biopsy and practical considerations. J Cutan Aesthet Surg. 2008 Jul;1(2):107-11. doi: 10.4103/0974-2077.44174. PMID 20300359
• Yasui Y, Kato H, Oda T, Nakamura M, Morita A. Complications and risk factors of punch biopsy: A retrospective large-scale study. J Dermatol. 2023 Jan;50(1):98-101. doi: 10.1111/1346-8138.16585. Epub 2022 Sep 24. PMID 36151785
• Akiyama Y, Norimatsu Y, Ohno Y. Prophylactic antimicrobials may not be needed to prevent surgical site infection after skin biopsy: a retrospective study. Antimicrob Resist Infect Control. 2022 Feb 16;11(1):35. doi: 10.1186/s13756-022-01077-z. PMID 35172906

Identifiers

NCT: NCT07596615 · MIRACLE · FIS-2024-00858

Primary sources (government registries)

• ClinicalTrials.gov (US NIH)

View this study on ClinicalTrials.gov ↗