Human menstrual blood-derived stem cells as immunoregulatory therapy in COVID-19: A case report and review of the literature.

BACKGROUND: The coronavirus disease 2019 (COVID-19) caused by novel coronavirus 2019 in December 2019 has spread all around the globe and has caused a pandemic. There is still no current effective guidance on the clinical management of COVID-19. Mesenchymal stem cell therapy has been shown to be one of the therapeutic approaches to alleviate pneumonia and symptoms through their immunomo-dulatory effect in COVID-19 patients. CASE
SUMMARY: We describe the first confirmed case of COVID-19 in Hangzhou to explore the role of human menstrual blood-derived stem cells (MenSCs) in the treatment of COVID-19. Moreover, we review the immunomodulation effect including non-specific and specific immune functions of MenSCs for the therapy of COVID-19.
CONCLUSION: MenSCs can be helpful to find a promising therapeutic approach for COVID-19. ©The Author(s) 2021. Published by Baishideng Publishing Group Inc. All rights reserved.

Core Tip: The coronavirus disease 2019 (COVID-19) is the word that certainly will not be forgotten by everybody who lives in the first half of the twenty-first century. It has led many researchers from different biomedical fields to find solutions or treatments to manage the pandemic. However, there is still no current effective guidance on the clinical management of COVID-19. Mesenchymal stem cells are widely used to treat tissue and organ injuries with effective immunomodulatory and repair capacities, which makes them ideal for allogenic adoptive transfer therapy. In this study, we describe the first confirmed case of COVID-19 in Hangzhou, China to explore the role of human menstrual blood-derived stem cells (MenSCs) in the treatment of COVID-19. Moreover, we review the immunomodulation effect including non-specific and specific immune functions of MenSCs for the therapy of COVID-19, which can be helpful to find a promising therapeutic approach for this disease.

INTRODUCTION

The outbreak of coronavirus disease 2019 (COVID-19) caused by novel coronavirus 2019 (2019-nCoV) in late December 2019 is increasing rapidly in an epidemic scale and has spread to over 200 countries[1,2]. It has rapidly transmitted and become a major concern all over the world. The epidemiology, clinical characteristics, and treatment of COVID-19 have been reported in many cases by many institutions[3]. However, there is still no current effective guidance from the World Health Organization on the clinical management of COVID-19[4].

Mesenchymal stem cells (MSCs) are widely used to treat tissue and organ injuries, with effective immunomodulatory and repair capacities and low immunogenicity. This makes them ideal for allogenic adoptive transfer therapy[5,6]. Human menstrual blood-derived stem cells (MenSCs) have become a promising alternative because they are easy to collect and isolate and do not involve ethical considerations[7,8]. Previously, our group revealed that MenSCs were effective for treating liver failure and lung injury[9,10]. Here, we present the first case of COVID-19 identified in Hangzhou, China, on January 19, 2020, and its treatment, including MenSC therapy, and review the immunoregulatory effect of MenSC in treatment of COVID-19.

CASE PRESENTATION

Chief complaints

A low-grade fever and fatigue accompanied by dizziness for 2 d.

History of present illness

On January 19, 2020, a 32-year-old man was referred to our hospital with a low-grade fever and fatigue accompanied by dizziness for 2 d. Given the COVID-19 outbreak in Wuhan and his symptoms, the local hospital immediately admitted him with suspected COVID-19. He was immediately placed in a quarantine ward and underwent examination. Sputum and throat swab specimens were collected at admission and tested by reverse transcription-polymerase chain reaction for SARS-Cov-2 RNA. The infection was confirmed the day of admission, in accordance with China Centers for Disease Control guidance.

History of past illness

He had no other underlying diseases. He worked and lived in Hangzhou, but had travelled to Wuhan for business one week earlier and contacted two colleagues there.

Personal and family history

There was no noteworthy personal or family medical history.

Physical examination

After hospitalization, the patient’s chief symptoms were an occasional cough, shortness of breath, and chest pain. On physical examination, he had short rough breaths. His temperature was elevated to 38.7 ºC, with an arterial oxygen tension (PaO2) of 82 mmHg under ambient air.

Laboratory examinations

Laboratory examinations showed normal leukocytes (9.3 × 109/L), neutrophils (6 × 109/L), and lymphocytes (2.3 × 109/L). The patient’s high-sensitivity C-reactive protein (hs-CRP) level was 10.8 mg/L. Laboratory testing showed sharply increased leukocytes (25.9 × 109/L) and neutrophils (23.2 × 109/L), and an hs-CRP level of 43.5 mg/L. On day 7, his fever reached 39.4 ºC and he developed chest tightness with a PaO2 of 68 mmHg, PaCO2 of 34 mmHg, and oxygenation index of 208 mmHg on nasal oxygen at 3 L/min. The levels of inflammatory cytokines were also increased, with interleukin (IL)-6 71.5 pg/mL, IL-10 7.75 pg/mL, tumor necrosis factor (TNF)-α 84.28 pg/mL, and TNF-γ 34.71 pg/mL on day 11. The sputum 2019-nCoV RNA test turned negative for the first time, although a repeat test was positive on day 20. The leukocyte count (14.6 × 109/L), neutrophil count (10.4 × 109/L), and inflammatory factor levels were all improved. Liver function tests showed elevated alanine aminotransferase (ALT, 190 U/L) and aspartate aminotransferase (AST, 41 U/L) levels on day 23.

Imaging examinations

The initial chest computed tomography (CT) scan showed a few interstitial changes in both lungs and ground-glass opacities (GGOs) in the subpleural area of the right lower lobe (Figure 1A). Subsequent chest CT showed multifocal peripheral patchy areas of nodular consolidation and new GGOs in the left subpleural area on day 7. Progressive resolution of the parenchymal lesions was seen on follow-up CT, which showed basilar streaky opacities and patchy consolidations, but worsened GGOs in both lungs (Figure 1B) on day 11. Chest CT on day 17 revealed improvement of the infiltrates in both lungs (Figure 1C). On day 24, the patient was discharged on glycyrrhizin tablets for liver protection. No abnormalities were observed on chest CT 9 d post-discharge (Figure 1D).

Figure 1

Chest computed tomography images of the patient with coronavirus disease 2019 in different stages of illness. A: Interstitial changes in both lungs and ground-glass opacities (GGOs) in the subpleural area of the right lower lobe on January 19 (Illness day 1); B: Worsening basilar streaky opacities, patchy consolidations, and GGOs in both lungs on January 30 (Illness day 12); C: Significant absorption of patches infiltrating in both lungs on February 4 (Illness day 17); D: Almost no abnormality with a few fibrotic changes left in both lungs on February 19.

MULTIDISCIPLINARY EXPERT CONSULTATION

Given the severe pulmonary injury caused by the inflammatory response and side effects, the glucocorticoid, antiviral, and antibiotic therapies were withdrawn. Under the guidance of a specialist group, MenSC therapy was proposed. The therapy was discussed and approved by the hospital ethics committee and the patient and family members provided informed consent before the therapy. Intravenous MenSCs were given at 3000, 2000, and 3000 U on hospital days 11, 12, and 14, respectively; no adverse events were observed in association with the infusion. Over the following 3 d, the patient’s breathing improved, with an intermittent dry cough and decreased chest stuffiness. His temperature decreased to 37.4 ºC.

FINAL DIAGNOSIS

The final diagnosis of the presented case was COVID-19 caused by 2019-nCoV.

TREATMENT

The patient was given supportive care including immunoglobulin and antiviral treatment at the first stage of hospitalization. Supportive treatment was strengthened, and methylprednisolone and vaccination were given as empirical treatment during the progression of the disease. Finally, MenSC therapy was proposed while the glucocorticoid, antiviral, and antibiotic therapies were withdrawn.

OUTCOME AND FOLLOW-UP

The condition of the patient was significantly improved. Regular follow-up revealed that no abnormalities or recurrences were observed on the chest CT after the patient’s discharge (Supplementary Figure 1).

DISCUSSION

This was the first confirmed case of COVID-19 in Hangzhou, Zhejiang Province, China. Most of the initial cases had a history of exposure in the epidemic area and were infected with the virus via human-to-human transmission[11,12]. The main clinical manifestations of COVID-19 infection are cough, fever, fatigue, and gradual dyspnea in some cases and acute respiratory distress syndrome in severe cases, with a 1–14-d period from onset to admission[13]. Our patient developed a mild fever and fatigue 4 d after contact in the epidemic area. As his illness worsened, he developed obvious respiratory symptoms, including a cough and chest discomfort. With treatment, his temperature normalized and the other complaints were relieved.

Our patient showed changes in routine laboratory parameters and inflammatory cytokines consistent with other reports[14]. The elevated ALT and AST levels observed late in the patient’s illness were thought to be adverse outcomes of systemic inflammation due to the virus[15]. Of note, the stool specimen collected on day 23 was negative for 2019-nCoV RNA, consistent with the sputum specimens[16].

Chest CT can help diagnose patients suspected of having COVID-19[17]. Similar to viral pneumonia due to other etiologies, GGOs are the main CT findings in COVID-19. The initial CT showed basilar streaky opacities and GGOs in the right lower lobe[18,19]. As the illness progressed, the size and density of these GGOs or paving patchy consolidations increased. Only slight fibrotic changes remained when the patient recovered.

The most effective medical treatment against COVID-19 remains unknown, though several antiviral therapies were confirmed to be effective in published studies[20]. Antibiotic was also adopted to prevent secondary bacterial infection in this case. Apart from the large supportive treatment, stem cell therapy was considered with no side effect after three times of intravenous administration. After the third administration, inflammatory cytokines such as IL-6 and IL-10 decreased significantly. Moreover, patchy consolidations were absorbed in both lungs, accompanied with the gradually well-improved clinical conditions.

The superiority of MSC therapy over therapies based on bone marrow, adipose, umbilical cord, or embryonic tissue is due to the easily accessible source of the cells and their high rate of proliferation, the low invasiveness of the procedure, and the absence of ethical issues[21,22]. MenSCs can be isolated from female uterine blood with readily accessible materials, free of trauma or ethical concerns, and are thus of great potential for treating diseases[23,24]. In vitro, MenSCs can be differentiated into ectoderm and mesoderm, specifically, into fat, bone, cartilage, nerve, and endothelioid cells[5,9]. The expression by MenSCs of OCT-4, specific embryonic antigen SSEA-4, C-kit, and other embryonic stem cell markers suggests that MenSCs are more primitive and have a stronger multidirectional differentiation potential than other MSCs[25-27]. Moreover, with their strong paracrine and angiogenic potential, they may be useful in the repair of damaged tissues[28].

MSCs interact with inhibitory T cells, B cells, natural killer (NK) cells, and regulatory T cells to regulate the immune response, which also have direct effects on NK cells[29-32]. Recent studies indicate that acute respiratory distress syndrome (ARDS), characterized by acute inflammation and edema of the alveolar epithelial cells, is accompanied by the massive release of inflammatory cytokines, such as IL-1, IL-8, interferon, and TNF, which activate macrophages or NK cells, which in a “cytokine storm” trigger a strong immune response to damage alveoli[23]. COVID-19 is associated with abundant complications, with ARDS as the main cause of death[33,34]. By inhibiting over-activated NK cells, MenSCs could confer protection against the highly inflammatory environment while maintaining the NK activities that reduce inflammation during healing. Moreover, immunoregulation by MenSCs via IL-6 and IL-10 has been reported as well[23,35,36].

MenSCs are significant inhibitors of the inflammatory response[37,38]. Their ability to inhibit T lymphocyte proliferation results in stronger immunomodulatory effects than those exhibited by human umbilical cord mesenchymal stem cells or bone marrow mesenchymal stem cells (BM-MSCs)[39,40]. The inhibition of T lymphocyte proliferation by MenSCs is mediated by their secretion of prostaglandin (PGE2) and indoleamine2, 3-dioxygenase (IDO)[40]. PGE2 contributes to the transformation of classically activated macrophages (M1) to replace activated macrophages (M2) and may therefore be of interest in ameliorating the over-active inflammatory response in COVID-19[41]. It has also been shown that MenSCs promote the proliferation of CD4+ T lymphocytes in a density-dependent manner that is not affected by the concentration of IDO, in contrast to BM-MSCs, which are sensitive to IDO[42].

In addition to interleukins or PGE2, MenSCs release hepatocyte growth factor, granulocyte-macrophage colony stimulating factor, and keratinocyte growth factor (KGF), which may protect respiratory epithelial cells in lung injury via activities that include the increased clearance of alveolar fluid, the promotion of endothelial repair, and the inhibition of the inflammatory response[10,43,44]. MenSCs cov-2downregulate caspase-3 and IL-1β expression and upregulate KGF expression to improve microvascular permeability. In turn, KGF activates alveolar type II epithelial cells to stimulate the synthesis of pulmonary surfactant[10,32,45].

These abilities of MenSCs suggest a capacity to regulate the inflammatory response and thereby overcome the severe cytokine storm associated with COVID-19, while restoring the normal function of immune cells and tissues. Based on their homing potential, MenSCs may enhance the repair properties of immune and other cells, thus allowing the reconstruction of damaged tissues by receptor-mediated interactions[46,47]. By protecting alveolar epithelial cells and improving the pulmonary microen-vironment, MSCs can contribute to preventing lung dysfunction and the development of COVID-19 pneumonia[48]. Through mediators such as interleukin receptors, PGE2, and KGF, MSCs can significantly inhibit the inflammatory response and repair the vascular-alveolar epithelial cell barrier to promote the elimination of alveolar fluid and thereby the infection[49,50].

CONCLUSION

This was the first COVID-19 case seen in Hangzhou, China. During the patient’s hospitalization, laboratory abnormalities and chest CT findings changed in synchrony with the clinical illness. The immune system of COVID-19 patients produces a drastic inflammatory response, causing a cytokine storm that includes the overproduction of cytokines or immune cells[51]. MSCs have been reported to enhance cellular reparative and immunomodulatory properties and thereby prevent or reduce the cytokine storm that develops in COVID-19 patients[52]. After the intravenous injection of MSCs, a portion of the cells accumulate in the lungs via the circulation, where they may contribute to the recovery of the pulmonary microenvironment by protecting alveolar epithelial cells, reducing the amount of effusion, and preventing pulmonary fibrosis, thus ameliorating tissue dysfunction and COVID-19 pneumonia[33]. Pulmonary fibrosis is a serious complication of COVID-19[53], with the first step in its pathogenesis consisting of the intra-alveolar infiltration of proliferating fibroblasts. Proliferating fibroblasts infiltrating intra-alveolar zone was considered the first step in pathogenesis of pulmonary fibrosis[54]. Edema, fibroblast invasion of the pulmonary interspaces, and type II pneumocyte hyperplasia with inflammatory cytokine infiltrations have been seen in the lung sections of patients with COVID-19[55]. MenSCs, alone or combined with other therapeutic agents, might be a promising treatment for COVID-19.

ACKNOWLEDGEMENTS

The authors would like to acknowledge Jiong Yu for their assistance in the conduct of the study.