Stephen E. Christmas

Mesenchymal stem cells (MSCs) modulate immune responses and maintain self-tolerance. Their trophic activities and regenerative properties make them potential immunosuppressants for treating autoimmune and autoinflammatory diseases. MSCs are drawn to sites of injury and inflammation where they can both reduce inflammation and contribute to tissue regeneration. An increased understanding of the role of MSCs in the development and progression of autoimmune disorders has revealed that MSCs are passive targets in the inflammatory process, becoming impaired by it and exhibiting loss of immunomodulatory activity. MSCs have been considered as potential novel cell therapies for severe autoimmune and autoinflammatory diseases, which at present have only disease modifying rather than curative treatment options. MSCs are emerging as potential therapies for severe autoimmune and autoinflammatory diseases. Clinical application of MSCs in rare cases of severe disease in which other existing treatment modalities have failed, have demonstrated potential use in treating multiple diseases, including rheumatoid arthritis, systemic lupus erythematosus, myocardial infarction, liver cirrhosis, spinal cord injury, multiple sclerosis, and COVID-19 pneumonia. This review explores the biological mechanisms behind the role of MSCs in autoimmune and autoinflammatory diseases. It also covers their immunomodulatory capabilities, potential therapeutic applications, and the challenges and risks associated with MSC therapy.

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Background: Therapeutic approaches for cancer rely on careful consideration of finding the optimal way of delivering the pro-drug for cellular-based cancer treatment. Cell lines and cell cultures have been used in these studies to compare the in vitro and in vivo efficacy of autologous vs. allogeneic tumour cellular gene therapy. Here we have investigated and are reporting for the first time the effect of prodrug ganciclovir (GCV)-preloading (pre-treatment) in suicide gene therapy of cancer. Methods: This study examines the effect of GCV-preloading (pre-treatment) on a range of tumour cell lines in conjunction with suicide gene therapy of cancer. To determine the efficacy of this modality, a series of in vitro and in vivo experiments were conducted using genetically modified and unmodified tumour cell lines. Results: Following co-culture of herpes simplex virus thymidine kinase (HSV-TK) modified tumour cells and unmodified tumour cells both in vitro and in vivo, GCV-preloading (pre-treatment) of TK-modified human and mouse mesothelioma cells and ovarian tumour cells allowed them to mediate efficiently bystander killing of neighbouring unmodified tumour cells in vitro. In contrast, GCV-preloading of TK-modified human and mouse mesothelioma cells and ovarian tumour cells abolished their in vivo ability to induce bystander killing of unmodified tumour cells, although there was some tumour regression compared to control groups but this was not statistically significant. These results suggest that preloading TK modified tumour cells with GCV needs further study to define the most effective strategy for an in vivo application to retain their bystander killing potential after exposure to lethal doses of GCV in vitro. Conclusions: This study highlights the promising possibility of improving the efficacy of pro-drug system to prevent any damage to the immune system and enhancing this type of suicide gene therapy of cancer, as well as the need for further studies to explore the discrepancies between in vitro and in vivo results.

Complement activation leads to membrane attack complex formation, which can lyse not only pathogens but also host cells. Histones can be released from the lysed or damaged cells and serve as a major type of damage-associated molecular pattern, but their effects on the complement system are not clear. In this study, we pulled down two major proteins from human serum using histone-conjugated beads: one was C-reactive protein and the other was C4, as identified by mass spectrometry. In surface plasmon resonance analysis, histone H3 and H4 showed stronger binding to C4 than other histones, with K_D around 1 nM. The interaction did not affect C4 cleavage to C4a and C4b. Because histones bind to C4b, a component of C3 and C5 convertases, their activities were significantly inhibited in the presence of histones. Although it is not clear whether the inhibition was achieved through blocking C3 and C5 convertase assembly or just through reducing their activity, the outcome was that both classical and mannose-binding lectin pathways were dramatically inhibited. Using a high concentration of C4 protein, histone-suppressed complement activity could not be fully restored, indicating C4 is not the only target of histones in those pathways. In contrast, the alternative pathway was almost spared, but the overall complement activity activated by zymosan was inhibited by histones. Therefore, we believe that histones inhibiting complement activation is a natural feedback mechanism to prevent the excessive injury of host cells.

Cellular based therapeutic approaches for cancer rely on careful consideration of finding the optimal cell to execute the cellular goal of cancer treatment. Cell lines and primary cell cultures have been used in some studies to compare the in vitro and in vivo efficacy of autologous vs allogeneic tumour cell vaccines. This study examines the effect of γ-irradiation on a range of tumor cell lines in conjunction with suicide gene therapy of cancer. To determine the efficacy of this modality, a series of in vitro and in vivo experiments were conducted using genetically modified and unmodified tumor cell lines. Following co-culture of HSV-TK modified tumor cells and unmodified tumor cells both in vitro and in vivo we observed that the PA-STK ovarian tumor cells were sensitive to γ-irradiation, completely abolishing their ability to induce bystander killing of unmodified tumor cells. In contrast, TK-modified human and mouse mesothelioma cells were found to retain their in vitro and in vivo bystander killing effect after γ-irradiation. Morphological evidence was consistent with the death of PA-STK cells being by pyknosis after γ-irradiation. These results suggest that PA-STK cells are not suitable for clinical application of suicide gene therapy of cancer, as lethal γ-irradiation (100 Gy) interferes with their bystander killing activity. However, the human mesothelioma cell line CRL-5830-TK retained its bystander killing potential after exposure to similarly lethal γ-irradiation (100 Gy). CRL-5830 may therefore be a suitable vehicle for HSV-TK suicide gene therapy. This study highlights the diversity among tumor cell lines and the careful considerations needed to find the optimal tumor cell line for this type of suicide gene therapy of cancer.

HLA‐G is a non‐classical class I HLA antigen, normally expressed in high levels only on extravillous cytotrophoblast. It has immunosuppressive properties in pregnancy and has also been found to be upregulated on leucocytes in viral infection. In this study, proportions of all leucocyte subsets expressing HLA‐G were found to be low in healthy subjects positive or negative for cytomegalovirus (CMV). Significantly greater proportions of CD4+ CD69+ and CD56+ T cells expressed HLA‐G compared to other T cells. However, following stimulation with CMV antigens or intact CMV, proportions of CD4+, CD8+, CD69+ and CD56+ T cells, and also B cells expressing HLA‐G, were significantly increased in CMV+ subjects. Despite some subjects having alleles of HLA‐G associated with high levels of expression, no relationship was found between HLA‐G genotype and expression levels. Purified B cells from CMV+ subjects stimulated in mixed culture with CMV antigens showed significantly increased HLA‐G mRNA expression by real‐time polymerase chain reaction. Serum levels of soluble HLA‐G were similar in CMV− and CMV+ subjects but levels in culture supernatants were significantly higher in cells from CMV+ than from CMV− subjects stimulated with CMV antigens. The HLA‐G ligand KIR2DL4 was mainly expressed on NK cells and CD56+ T cells with no differences between CMV+ and CMV− subjects. Following stimulation with IL‐2, an increase in the proportion of CD56+ T cells positive for KIR2DL4 was found, together with a significant decrease in CD56dimCD16+ NK cells. The results show that CMV influences HLA‐G expression in healthy subjects and may contribute to viral immune evasion.

Killer Cell Immunoglobulin-like Receptors (KIR) have been used as good markers for the study of genetic predisposition in many diseases and in human genetic population dynamics. In this context, we have investigated the genetic diversity of KIR genes and their main HLA class I ligands in Saudi population and compared the data with other studies of neighboring populations. One hundred and fourteen randomly selected healthy Saudi subjects were genotyped for the presence or absence of 16 KIR genes and their HLA-C1, -C2, -Bw4(Thr80) and Bw4(Ile80) groups, using a PCR-SSP technique. The results show the occurrence of the framework genes (3DL2, 3DL3 and 2DL4) and the pseudogenes (2DP1 and 3DP1) at highest frequencies. All inhibitory KIR (iKIR) genes appeared at higher frequencies than activating genes (aKIR), except for 2DS4 with a frequency of 90.35%. A total of 55 different genotypes were observed appearing at different frequencies, where 12 are considered novel. Two haplotypes were characterized, AA and Bx (BB and AB), which were observed in 24.5% and 75.5% respectively of the studied group. The frequencies of iKIR + HLA associations were found to be much higher than aKIR + HLA. KIR genes frequencies in the Saudi population are comparable with other Middle Eastern and North African populations.

Aims: We performed an association study to evaluate the contribution of 16 killer cell immunoglobulin-like receptor (KIR) genotype polymorphisms and the HLA-C1 and -C2 ligands in the development of colorectal cancer (CRC) in Saudi Arabian patients. Methods: A total of 52 patients with different stages of malignant CRC as well as 70 healthy Saudi controls were enrolled at the King Khalid University Hospital. Results: Our results showed that the frequency of the activating mutations KIR2DS1, 2DS2, 2DS3, 2DS5, and 3DS1 was significantly higher in CRC patients compared to controls. The 3DS1 gene contributed to the highest risk of CRC (odds ratio [OR] = 16.25, p < 0.0001), followed by 2DS1 (OR = 8.6; p < 0.0001). The distributions of HLA-C1 and -C2 ligands were not significantly different between patients and controls. Analyses of different combinations of KIR genes with their HLA-C1 and -C2 ligands show that the frequency of 2DL3 in the presence of its ligand, the allotype C1, was significantly more prevalent in patients compared to controls. In addition, 2DL2 and 2DL3 that were aggregated in combination with the ligand, HLA-C1, were found to be more highly associated mainly with the homozygote HLA-C1/C1 (p = 0.03; OR = 2.6). The activating mutations 2DS1 and 2DS2 when combined with their respective ligands, HLA-C2 and -C1, showed highly significant associations with CRC development. Conclusion: This study supports a key role for KIR gene mutations in the development of CRC, especially in association with their ligands.

Background: CD56+ T cells previously have been identified as potentially cytotoxic lymphocytes, and relative numbers are increased in some infectious diseases.Patients and methods: Relative proportions ofCD56+ T cells were measured by flow cytometry in groups of renal transplant patients differing in cytomegalovirus (CMV) status of donor (D) and recipient (R). These measurements were related to episodes ofCMVviremia.Results: Patient groups in which recipients (R+) or donors (D+/R−) wereCMV+ had significantly higher proportions ofCD56+ T cells (5.11 ± 0.69% and 5.42 ± 1.01%, respectively) than the D−/R− group (1.9 ± 0.35%;P = 0.0018 and 0.017, respectively). In the high‐risk D+/R− group, it was found that patients who had post‐transplantCMVviremia had higher levels than those who remainedCMVnegative (9.09 ± 2.34% vs. 3.16 ± 1.22%;P = 0.01).CD56+ T cells from R+ and D+/R− groups had higher proportions of bothCD4+ andCD8+ cells than the D−/R− group. When activation markers were examined, someCD56+ T cells from bothCMV+ groups had a T_EMphenotype, with significantly more expressingCD45ROandNKG2C, and less expressingCD28,CD62L,CD127, andCD161 compared to the D−/R− group. SomeCD56+ T cells showed specificity forCMVantigens and similar proportions ofCD8+ cells were positive for class IHLA‐CMVtetramers containing immunodominantCMVpeptides compared to the majorityCD56− T cells.Conclusion: The results show significant increases in proportions ofCD56+ T cells in relation toCMVinfection in renal transplant patients and suggest that these cells have a cytotoxic function againstCMV‐infected cells.