Effect of COVID-19 on Platelet Mitochondrial Bioenergetic, Antioxidants and Oxidative Stress in Infertile Men.

Overview

To verify the hypothesis that infertility and the effect of SARS-CoV-2 on infertility may damage platelet mitochondrial bioenergetics and endogenous coenzyme Q10 levels in infertile men.

Full Title of Study: “Effect of COVID-19 on Spermiogram, Platelet Mitochondrial Bioenergetic, Antioxidants and Oxidative Stress in Infertile Men”

Study Type

• Study Type: Interventional
• Study Design
  • Allocation: Randomized
  • Intervention Model: Parallel Assignment
  • Primary Purpose: Other
  • Masking: None (Open Label)
• Study Primary Completion Date: December 31, 2022

Detailed Description

Infertility is defined as the failure of the reproductive system to achieve pregnancy after 12 months of unprotected sex life. The pathobiochemical mechanisms of male fertility disorders include reduced sperm motility and quality, oxidative stress, reduced antioxidant capacity, mtDNA fragmentation, and sperm mitochondrial dysfunction. Sperm contain a number of mitochondria that are spirally arranged around the middle part of the axomen. The main role of mitochondria in spermatozoa is to generate the energy needed for their motility (1, 2). Endogenous sources – coenzyme Q10 and carnitine – are key for energy production (ATP) in sperm mitochondria. Physiological functions of sperm require a minimal amount of reactive oxygen species (ROS), but uncontrolled ROS production contributes to reduced motility and sperm count, fragmentation of mtDNA (3). In recent years, blood cells (platelets, lymphocytes and monocytes) have been used to diagnose mitochondrial disorders. Isolated peripheral blood platelets are an available source of mitochondria to assess mitochondrial health. Platelets receive energy mainly through glycolysis and oxidative phosphorylation. Platelet mitochondrial dysfunction has been demonstrated in patients with chronic kidney disease (4, 5), in patients with rheumatoid arthritis (6), in patients with acute COVID-19 (7). An O2k-respirometer (Oroboros, Austria) (8, 9) is used for respirometric analysis of platelet mitochondrial bioenergetics. None information is available on the effect of infertility on platelet mitochondrial function, none on the effect of SARS-CoV-2 on platelet mitochondrial function in infertile patients, or the effect of vaccination on sperm function. Testicular damage and subsequent infertility due to SARS-CoV infection is expected. -2, directly via SARS-CoV-2 binding to ACE2 receptors or secondarily, in relation to the immunological and inflammatory response (10). SARS-CoV-2 virus induces excessive production of pro-inflammatory cytokines, mainly interleukin 6 (IL6), interleukin-1β (IL-1β) and tumor necrosis factor α (TNFα). Cytokines can impair sperm movement and reduce sperm count. High levels of pro-inflammatory cytokines have been found in infertile men (with oligozoospermia, asthenozoospermia, teratozoospermia) (11). SARS-CoV-2 virus can manipulate mitochondrial function in patients with post-COVID-19 syndrome, which may persist for a long time (12). In previous our study the investigators found modulation of platelet mitochondrial respiration, reduction ATP production via oxidative phosphorylation, reduces endogenous coenzyme Q10 production, reprogramming of cellular metabolism patients after 4-7 weeks overcoming acute COVID-19, SARS-CoV-2 (7). In another studies the investigators confirmed platelet mitochondrial bioenergetic deficiency, reduced endogenous coenzyme Q10 production in patients with post-COVID-19 syndrome, 3-6 months after overcoming COVID-19 (13, 14, 15). Results of this study contribute to the understanding of the pathobiochemical mechanisms of infertility on subcellular level and to verify the hypothesis that infertility and the effect of SARS-CoV-2 on infertility may affect platelet mitochondrial bioenergetics and endogenous coenzyme Q10 levels.

Interventions

• Other: diagnostic test and sperm analysis
  • Diagnostic Test: 2×14 ml of peripheral blood collected in the tube with anticoagulant, for platelet isolation, respirometry mitochondrial analysis, antioxidants (coenzyme Q10, vitamin E, gamma-tocopherol, beta-carotene) and TBARS estimation. Sperm analysis: standard spermiogram examination (volume, pH, number, motility and pathology) in the broker chamber, as well as extended examinations: mioxsys for redox potential, Vitalsperm (eosin-nigrosine staining) for sperm vitality and anti-sperm antibody (IgG) test.

Arms, Groups and Cohorts

• Active Comparator: infertile men with post-COVID-19 (vaccinated or without vaccination)
  • 15 infertile men with post-COVID-19 (vaccinated or without vaccination)
• Active Comparator: infertile men without post-COVID-19 (vaccinated or without vaccination)
  • 15 infertile men without post-COVID-19 (vaccinated or without vaccination)
• Active Comparator: Control: 15 healthy men volunteers (no COVID-19, no other pathologies)
  • 15 healthy men volunteers (no COVID-19, no other pathologies) as control group

Clinical Trial Outcome Measures

Primary Measures

• Clinical symptoms
  • Time Frame: 15 minutes
  • Clinical symptoms: infertile patients without COVID-19 (vaccinated, or none vaccinated) Clinical symptoms patients after COVID-19
• Damaged platelet mitochondrial bioenergetics 1
  • Time Frame: 1 day
  • Basal oxygen consumption rate in intact platelets (ce)
• Damaged platelet mitochondrial bioenergetics 2
  • Time Frame: 1 day
  • rate of mitochondria LEAK respiration with CI-linked substrates (1PM – state 4)
• Damaged platelet mitochondrial bioenergetics 3
  • Time Frame: 1 day
  • CI-linked respiration coupled with ATPproduction (2D-CI-linked oxidative phosphorylation capacity), respiration after addition of cytochrome c (2C).
• Damaged platelet mitochondrial bioenergetics 4
  • Time Frame: 1 day
  • Maximal oxidative capacity (the electron transfer capacity -ET), after uncoupler titration (3U).
• Damaged platelet mitochondrial bioenergetics 5
  • Time Frame: 1 day
  • After addition of exogenous substrate glutamate (4G) non-coupled mitochondrial oxygen consumption.
• Damaged platelet mitochondrial bioenergetics 6
  • Time Frame: 1 day
  • Non-coupled oxygen consumption with CI&CII-linked substrate (5S) improvement of mitochondrial parameters representing OXPHOS- and electron tranport capacity (ET-capacity).
• Endogenous coenzyme Q10-TOTAL 1
  • Time Frame: 1 day
  • CoQ10-TOTAL in: Platelets (pmol.10-9 cells)
• Endogenous coenzyme Q10-TOTAL 2
  • Time Frame: 1 day
  • CoQ10-TOTAL in: Blood (µmol.L-1)
• Endogenous coenzyme Q10-TOTAL 3
  • Time Frame: 1 day
  • CoQ10-TOTAL in: Plasma (µmol.L-1)
• Endogenous coenzyme TBARS
  • Time Frame: 1 day
  • Endogenous concentration of CoQ10-TOTAL (ubiquinone + ubiquinol) in platelets, blood and plasma CoQ10-TOTAL in: TBARS in plasma (µmol.L-1).
• Sperm analysis 1
  • Time Frame: 1 day
  • standard spermiogram examination (volume, pH, number, motility and pathology) in the broker chamber
• Sperm analysis 2
  • Time Frame: 1 day
  • mioxsys for redox potential
• Sperm analysis 3
  • Time Frame: 1 day
  • Vitalsperm (eosin-nigrosine staining) for sperm vitality
• Sperm analysis 4
  • Time Frame: 1 day
  • anti-sperm antibody (IgG) test

Participating in This Clinical Trial

Inclusion Criteria

• Infertile patients without COVID-19 – Infertile patients after COVID-19 Control group: healthy volunteers Exclusion Criteria:

• disagreement with informed consent

Gender Eligibility: Male

Only men have sperm.

Minimum Age: 18 Years

Maximum Age: 40 Years

Are Healthy Volunteers Accepted: Accepts Healthy Volunteers

Investigator Details

• Lead Sponsor
  • Comenius University
• Collaborator
  • GYN-FIV
• Provider of Information About this Clinical Study
  • Sponsor
• Overall Official(s)
  • Anna Gvozdjáková, Prof.Dr.DSc., Principal Investigator, CU in Bratislava, Faculty of Medicine, Pharmacobiochemical Laboratory of 3rd department of Medicine

Citations Reporting on Results

Sheweita SA, Tilmisany AM, Al-Sawaf H. Mechanisms of male infertility: role of antioxidants. Curr Drug Metab. 2005 Oct;6(5):495-501. doi: 10.2174/138920005774330594.

Gvozdjakova A, Kucharska J, Dubravicky J, Mojto V, Singh RB. Coenzyme Q(1)(0), alpha-tocopherol, and oxidative stress could be important metabolic biomarkers of male infertility. Dis Markers. 2015;2015:827941. doi: 10.1155/2015/827941. Epub 2015 Feb 25.

Gvozdjakova A, Sumbalova Z, Kucharska J, Chladekova A, Rausova Z, Vancova O, Komlosi M, Ulicna O, Mojto V. Platelet mitochondrial bioenergetic analysis in patients with nephropathies and non-communicable diseases: a new method. Bratisl Lek Listy. 2019;120(9):630-635. doi: 10.4149/BLL_2019_104.

Gvozdjakova A, Sumbalova Z, Kucharska J, Komlosi M, Rausova Z, Vancova O, Szamosova M, Mojto V. Platelet Mitochondrial Respiration, Endogenous Coenzyme Q10 and Oxidative Stress in Patients with Chronic Kidney Disease. Diagnostics (Basel). 2020 Mar 23;10(3):176. doi: 10.3390/diagnostics10030176.

Gvozdjakova A, Sumbalova Z, Kucharska J, Szamosova M, Capova L, Rausova Z, Vancova O, Mojto V, Langsjoen P, Palacka P. Platelet mitochondrial respiration and coenzyme Q10 could be used as new diagnostic strategy for mitochondrial dysfunction in rheumatoid diseases. PLoS One. 2021 Sep 28;16(9):e0256135. doi: 10.1371/journal.pone.0256135. eCollection 2021.

Sumbalova Z, Kucharska J, Palacka P, Rausova Z, Langsjoen PH, Langsjoen AM, Gvozdjakova A. Platelet mitochondrial function and endogenous coenzyme Q10 levels are reduced in patients after COVID-19. Bratisl Lek Listy. 2022;123(1):9-15. doi: 10.4149/BLL_2022_002.

Pesta D, Gnaiger E. High-resolution respirometry: OXPHOS protocols for human cells and permeabilized fibers from small biopsies of human muscle. Methods Mol Biol. 2012;810:25-58. doi: 10.1007/978-1-61779-382-0_3.

Abobaker A, Raba AA. Does COVID-19 affect male fertility? World J Urol. 2021 Mar;39(3):975-976. doi: 10.1007/s00345-020-03208-w. Epub 2020 Apr 21. No abstract available.

Rajak P, Roy S, Dutta M, Podder S, Sarkar S, Ganguly A, Mandi M, Khatun S. Understanding the cross-talk between mediators of infertility and COVID-19. Reprod Biol. 2021 Dec;21(4):100559. doi: 10.1016/j.repbio.2021.100559. Epub 2021 Sep 1.

Gvozdjakova A, Klauco F, Kucharska J, Sumbalova Z. Is mitochondrial bioenergetics and coenzyme Q10 the target of a virus causing COVID-19? Bratisl Lek Listy. 2020;121(11):775-778. doi: 10.4149/BLL_2020_126.

Boguenet M, Bouet PE, Spiers A, Reynier P, May-Panloup P. Mitochondria: their role in spermatozoa and in male infertility. Hum Reprod Update. 2021 Jun 22;27(4):697-719. doi: 10.1093/humupd/dmab001.

Khalili MA, Leisegang K, Majzoub A, Finelli R, Panner Selvam MK, Henkel R, Mojgan M, Agarwal A. Male Fertility and the COVID-19 Pandemic: Systematic Review of the Literature. World J Mens Health. 2020 Oct;38(4):506-520. doi: 10.5534/wjmh.200134. Epub 2020 Aug 14.