Publikationsdatenbank

Susceptibility of humanized mice to Staphylococcus aureus in a localized deep-tissue abscess model (2023)

Art

Hochschulschrift

Autor

Hung, Sophia Isabella Gertrud (WE 7)

Quelle

Berlin, 2023 — XI, 95 Seiten

Sprache

Englisch

Verweise

URL (Volltext): https://refubium.fu-berlin.de/handle/fub188/41500

Kontakt

Institut für Mikrobiologie und Tierseuchen

Robert-von-Ostertag-Str. 7-13
14163 Berlin
+49 30 838 51843 / 66949
mikrobiologie@vetmed.fu-berlin.de

Abstract / Zusammenfassung

Staphylococcus aureus is a widespread Gram-positive bacterium that colonizes up to 80% of the human population as a facultative pathogen. It can cause a variety of diseases, ranging from mild skin infections to life-threatening endocarditis, pneumonia, and sepsis. S. aureus is particularly notorious for its ability to develop resistances against antimicrobial agents, a process that dates back to the discovery of penicillin. Today, methicillin-resistant S. aureus presents one of the greatest burdens to our healthcare system. The development of new treatments is therefore of the utmost urgency, especially that of an effective vaccine. So far, however, all promising candidates have failed as soon as they reached the clinical phase. One of the main reasons for this is the species-specificity of S. aureus, which is equipped with several virulence factors that are adapted to the human immune system. A promising new method for bridging this translational gap are humanized mice. These are immunodeficient mouse strains engrafted with human stem cells, leading to the development of a viable human immune system. One of the most commonly used strains is the NSG mouse, which was also chosen in the S. aureus infection studies conducted so far. These encompass only a small number of models, including peritonitis, skin infection, pneumonia, and osteomyelitis. Humanized NSG (huNSG) mice were more susceptible to the pathogen than the control groups in all of these models. For this thesis, 6 to 8 week old NSG mice were engrafted with cord-blood derived human stem cells and infected after 18 weeks with S. aureus in a localized, deep-tissue abscess model. The huNSG mice and the control groups, wild-type mice (Balb/c), murinized NSG mice (mice implanted with murine bone marrow cells), and non-engrafted NSG mice were weighed every day and their general condition was checked. In addition, the bioluminescence signal in the infected muscle was measured daily. The animals were randomly removed from the experiment after two or seven days, to cover both the acute and the chronic stages of the infection. The huNSG mice consistently showed stronger bioluminescence signals and a higher bacterial burden in the muscle could be detected on day 7. The difference in survival between the groups was particularly striking. All control animals (except one non-engrafted NSG mouse) assigned to the day-7 group survived until the end of the experiment. In contrast, 7 of 20 huNSG animals reached the humane endpoint before. In huNSG mice, the bacteria spread to significantly more inner organs than in the wild-type and the murinized mice, where they also reached a higher bacterial count. Importantly, a direct correlation between the survival probability and the degree of humanization could be determined. HuNSG mice had significantly higher human and murine cytokine levels in the blood and the muscle than the control groups. The dynamics between the human and murine cytokines also varied greatly. While the levels of murine cytokines remained mostly constant, their human counterparts decreased significantly between day 2 and day 7. The number of human B cells also decreased noticeably in the bone marrow, the same being true for monocytes in the blood and bone marrow. In contrast, murine monocytes increased significantly. Humanized NSG mice sustain only very small numbers of human neutrophil cells, which play an important role in the human immune defence against S. aureus. Consequently, humanized NSG-SGM3 (huSGM3) animals were used for the second trial, as this mouse strain is equipped with three human cytokines that support the formation of myeloid cell lineages. However, overexpression of these cytokines also leads to undesirable side effects in these mice. The animals become anaemic during long-term humanization and eventually die. For this reason, the huSGM3 mice entered the experiments 12 weeks post engraftment. At this time, the animals showed a robust degree of humanization and only slightly decreased erythrocyte and haemoglobin levels. The humanized SGM3 mice were significantly more sensitive in the muscle abscess model than the control groups, including the HuNSG mice. The mice reached the humane endpoint earlier and in such large numbers, that not enough animals were available for the day 7 group. No differences could be detected with respect to the bacterial load in muscle and internal organs that would have explained the death of the animals. In addition, the number of murine immune cells available did not vary between the two humanized groups. In contrast, the huSGM3 mice had significantly more human cells in the blood and spleen; these included B cells, T cells, monocytes, and neutrophils. Individual cytokine levels, namely hCCL-2, hIL-8, hIL-6, and hTNF-α, were also significantly increased in the humanized SGM3 mice. Similar to the first trial, a direct correlation between the degree of humanization and a severe progression of the infection could be found. Further experiments are needed to fully understand the interactions between the bacterium and the human immune system in the humanized mice. It will be particularly important to characterize the role of virulence factors, such as the Panton-Valentin-Leukocidin, which are characterized by their affinity to human cells. Nevertheless, the results obtained in this thesis represent important new gains in knowledge, which may contribute to the development of an effective vaccine against S. aureus.