In vitro model of HIV virus production in postmortem leucocytes identified with HIV RNA in culture

Journals

Title：
In vitro model of HIV virus production in postmortem leucocytes identified with HIV RNA in culture
Author：
Klarisa¹^{, *}, Budi Sampurna, Aria Yudhistira¹, Djaja Surya Atmaja¹, Budiman Bela²
Author Affiliation：
1.Department of Forensic Science and Medicolegal, Dr. Cipto Mangunkusumo National General Hospital, Jakarta, Indonesia.
2.Department of Microbiology, Dr. Cipto Mangunkusumo National General Hospital, Jakarta, Indonesia
Received：Aug.6, 2023
Accepted：Sep.18, 2023
Published：Sep.22, 2023

Abstract

Purpose: The research aimed to investigate in vitro HIV replication in leucocytes imitating dead bodies' physiological oxygen deprivation condition. Methods: The research was conducted using experimental design. Blood samples were obtained from living persons infected by HIV to substitute for blood from dead bodies. Blood samples were studied for 48 hours at a temperature of 26-32°C with oxygen deprivation as a post-mortem condition. The plasma of HIV-infected blood was examined for viral load. The leucocytes were separated from the red blood cells and mixed with a plasma of HIV-positive individuals. This suspension was stored in tubes and heated cooking oil was overlayed on the surface of the plasma and leucocyte suspension to prevent oxygen exposure. Following centrifugation of suspension, the leucocytes were cultured. RT PCR was employed to detect virus production in the culture supernatant. Results: Polyacrylamide gel analysis showed that new RNA HIV was detected by RT PCR up to 48 hours of oxygen deprivation. This study also found morphologic changes in leucocytes due to cytopathic effects in the form of cell-to-cell infections. Conclusion: HIV can still replicate and produce new viruses after the isolation of leucocytes from a living person’s blood with oxygen deprivation.

Keywords

Blood culture, HIV in human postmortem/dead body, HIV replication, HIV RT PCR, Oxygen deprivation.

Doi

https://doi.org/10.58396/bephs020103

References

[1] K. Okaba et al., “Prevalence of blood-borne infections in forensic samples: Epidemiology in areas of Chiba, Japan.,” J. Forensic Leg. Med., vol. 90, p. 102396, Aug. 2022, doi: 10.1016/j.jflm.2022.102396.

[2] J. L. Burton, “Health and safety at necropsy.,” J. Clin. Pathol., vol. 56, no. 4, pp. 254–260, Apr. 2003, doi: 10.1136/jcp.56.4.254.

[3] Z. Ivanovic, “Physiological, ex vivo cell oxygenation is necessary for a true insight into cytokine biology.,” Eur. Cytokine Netw., vol. 20, no. 1, pp. 7–9, Mar. 2009, doi: 10.1684/ecn.2009.0144.

[4] J. M. Costin, “Cytopathic mechanisms of HIV-1.,” Virol. J., vol. 4, p. 100, Oct. 2007, doi: 10.1186/1743-422X-4-100.

[5] M. T. Liu et al., “Study on molecular mechanisms of CD4 dependency and independency of HIV-1 gp120,” RSC Adv., vol. 13, no. 9, pp. 6274–6286, 2023, doi: 10.1039/d3ra00433c.

[6] H. Leroy, M. Han, M. Woottum, L. Bracq, M. Xie, and S. Benichou, “Virus-Mediated Cell-Cell Fusion,” pp. 1–28.

[7] F. Hasan, Y. Chiu, R. M. Shaw, J. Wang, and C. Yee, “Hypoxia acts as an environmental cue for the human tissue-resident memory T cell differentiation program.,” JCI insight, vol. 6, no. 10, May 2021, doi: 10.1172/jci.insight.138970.

[8] H. Dokgöz, N. Arican, I. Elmas, and S. K. Fincanci, “Comparison of morphological changes in white blood cells after death andin vitro storage of blood for the estimation of postmortem interval.,” Forensic Sci. Int., vol. 124, no. 1, pp. 25–31, Dec. 2001, doi: 10.1016/s0379-0738(01)00559-x.

[9] H. Gill-King, Forensic Taphonomy: The Postmortem Fate of Human Remains. CRC Press LLC, 1997.

[10] C. J. Babapulle and N. P. Jayasundera, “Cellular changes and time since death.,” Med. Sci. Law, vol. 33, no. 3, pp. 213–222, Jul. 1993, doi: 10.1177/002580249303300306.

[11] P. Li et al., “Initiation of reverse transcription during cell-to-cell transmission of human immunodeficiency virus infection uses pre-existing reverse transcriptase.,” J. Gen. Virol., vol. 75 (Pt 8), pp. 1917–1926, Aug. 1994, doi: 10.1099/0022-1317-75-8-1917.

[12] T. Kok, P. Li, and C. Burrell, “Cell-to-cell transmission of human immunodeficiency virus infection induces two distinct phases of viral RNA expression under separate regulatory control.,” J. Gen. Virol., vol. 74 (Pt 1), pp. 33–38, Jan. 1993, doi: 10.1099/0022-1317-74-1-33.

[13] J. D. Lifson et al., “Induction of CD4-dependent cell fusion by the HTLV-III/LAV envelope glycoprotein.,” Nature, vol. 323, no. 6090, pp. 725–728, Oct. 1986, doi: 10.1038/323725a0.

[14] R. Leonard, D. Zagury, I. Desportes, J. Bernard, J. F. Zagury, and R. C. Gallo, “Cytopathic effect of human immunodeficiency virus in T4 cells is linked to the last stage of virus infection.,” Proc. Natl. Acad. Sci. U. S. A., vol. 85, no. 10, pp. 3570–3574, May 1988, doi: 10.1073/pnas.85.10.3570.

[15] M. Brioschi et al., “A mass spectrometry-based workflow for the proteomic analysis of in vitro cultured cell subsets isolated by means of laser capture microdissection.,” Anal. Bioanal. Chem., vol. 406, no. 12, pp. 2817–2825, May 2014, doi: 10.1007/s00216-014-7724-9.

[16] S. Eligini et al., “Human monocyte-derived macrophages spontaneously differentiated in vitro show distinct phenotypes.,” J. Cell. Physiol., vol. 228, no. 7, pp. 1464–1472, Jul. 2013, doi: 10.1002/jcp.24301.

[17] H. G. Drexler and C. C. Uphoff, “Mycoplasma contamination of cell cultures: Incidence, sources, effects, detection, elimination, prevention.,” Cytotechnology, vol. 39, no. 2, pp. 75–90, Jul. 2002, doi: 10.1023/A:1022913015916.

[18] K. Dennert and R. Kumar, “Traceability Methods for Cell Line Authentication and Mycoplasma Detection.,” SLAS Technol., vol. 26, no. 6, pp. 630–636, Dec. 2021, doi: 10.1177/24726303211030290.

[19] J. Ni, D. Wang, and S. Wang, “The CCR5-Delta32 Genetic Polymorphism and HIV-1 Infection Susceptibility: a Meta-analysis.,” Open Med. (Warsaw, Poland), vol. 13, pp. 467–474, 2018, doi: 10.1515/med-2018-0062.

[20] N. Totani, M. Yawata, T. Mori, and E. G. Hammond, “Oxygen content and oxidation in frying oil.,” J. Oleo Sci., vol. 62, no. 12, pp. 989–995, 2013, doi: 10.5650/jos.62.989.

[21] N. Totani, S. Tateishi, T. Mori, and E. G. Hammond, “Oxidation of frying oils during intermittent usage.,” J. Oleo Sci., vol. 61, no. 11, pp. 601–607, 2012, doi: 10.5650/jos.61.601.