Distinctive Dynamics and Functions of the CD4+CD25+FOXP3+ Regulatory T Cell Population in Patients with Severe and Mild COVID-19.

Although CD4+CD25+FOXP3+ regulatory T (TREG) cells have been studied in patients with COVID-19, changes in the TREG cell population have not been longitudinally examined during the course of COVID-19. In this study, we longitudinally investigated the quantitative and qualitative changes in the TREG cell population in patients with COVID-19. We found that the frequencies of total TREG cells and CD45RA-FOXP3hi activated TREG cells were significantly increased 15-28 d postsymptom onset in severe patients, but not in mild patients. TREG cells from severe patients exhibited not only increased proliferation but also enhanced apoptosis, suggesting functional derangement of the TREG cell population during severe COVID-19. The suppressive functions of the TREG cell population did not differ between patients with severe versus mild COVID-19. The frequency of TREG cells inversely correlated with SARS-CoV-2-specific cytokine production by CD4+ T cells and their polyfunctionality in patients with mild disease, suggesting that TREG cells are major regulators of virus-specific CD4+ T cell responses during mild COVID-19. However, such correlations were not observed in patients with severe disease. Thus, in this study, we describe distinctive changes in the TREG cell population in patients with severe and mild COVID-19. Our study provides a deep understanding of host immune responses upon SARS-CoV-2 infection in regard to TREG cells.

Duration of Replication-Competent Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Shedding Among Patients With Severe or Critical Coronavirus Disease 2019 (COVID-19).

BACKGROUND: Patterns of shedding replication-competent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in severe or critical COVID-19 are not well characterized. We investigated the duration of replication-competent SARS-CoV-2 shedding in upper and lower airway specimens from patients with severe or critical coronavirus disease 2019 (COVID-19). METHODS: We enrolled patients with active or recent severe or critical COVID-19 who were admitted to a tertiary care hospital intensive care unit (ICU) or long-term acute care hospital (LTACH) because of COVID-19. Respiratory specimens were collected at predefined intervals and tested for SARS-CoV-2 using viral culture and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Clinical and epidemiologic metadata were reviewed. RESULTS: We collected 529 respiratory specimens from 78 patients. Replication-competent virus was detected in 4 of 11 (36.3%) immunocompromised patients up to 45 days after symptom onset and in 1 of 67 (1.5%) immunocompetent patients 10 days after symptom onset (P = .001). All culture-positive patients were in the ICU cohort and had persistent or recurrent symptoms of COVID-19. Median time from symptom onset to first specimen collection was 15 days (range, 6-45) for ICU patients and 58.5 days (range, 34-139) for LTACH patients. SARS-CoV-2 RNA was detected in 40 of 50 (80%) ICU patients and 7 of 28 (25%) LTACH patients. CONCLUSIONS: Immunocompromise and persistent or recurrent symptoms were associated with shedding of replication-competent SARS-CoV-2, supporting the need for improving respiratory symptoms in addition to time as criteria for discontinuation of transmission-based precautions. Our results suggest that the period of potential infectiousness among immunocompetent patients with severe or critical COVID-19 may be similar to that reported for patients with milder disease.

CD39+ tissue-resident memory CD8+ T cells with a clonal overlap across compartments mediate antitumor immunity in breast cancer.

Despite being a standard treatment option in breast cancer, immune checkpoint inhibitors (ICIs) are only efficacious for a subset of patients. To gain a better understanding of the antitumor immune response in breast cancer, we examined the heterogeneity of CD8+ T cells in tumors, metastatic lymph nodes (mLNs), and peripheral blood from patients with early breast cancer (n = 131). Among tissue-resident memory CD8+ T (TRM) cells, including virus- and tumor-specific CD8+ T cells, CD39 expression was observed in a tumor-specific and exhausted subpopulation in both tumors and mLNs. CD39+ TRM cells from tumors and mLNs exhibited a phenotypic similarity and clonally overlapped with each other. Moreover, tumor or mLN CD39+ TRM cells clonally overlapped with CD39- TRM and non-TRM cells in the same compartment, implying a tissue-specific differentiation process. These inter-subpopulationally overlapping CD39+ TRM clonotypes were frequently detected among effector memory CD8+ T cells in peripheral blood, suggesting a systemic clonal overlap. CD39+ TRM cell enrichment was heterogeneous among molecular subtypes of breast cancer, which is associated with the different role of antitumor immune responses in each subtype. In vitro blockade of PD-1 and/or CTLA-4 effectively restored proliferation of CD39+ TRM cells and enhanced cytokine production by CD8+ T cells from tumors or mLNs, particularly in the presence of CD39+ TRM enrichment. This suggests that CD39+ TRM cells have a capacity for functional restoration upon ICI treatment. Thus, our study indicates that CD39+ TRM cells with a clonal overlap across compartments are key players in antitumor immunity in breast cancer.

The generation of stem cell-like memory cells early after BNT162b2 vaccination is associated with durability of memory CD8+ T cell responses.

COVID-19 vaccines elicit humoral and cellular immune responses. Durable maintenance of vaccine-induced immunity is required for long-term protection of the host. Here, we examine activation and differentiation of vaccine-induced CD8+ T cells using MHC class I (MHC-I) multimers and correlations between early differentiation and the durability of CD8+ T cell responses among healthcare workers immunized with two doses of BNT162b2. The frequency of MHC-I multimer+ cells is robustly increased by BNT162b2 but decreases 6 months post-second vaccination to 2.4%-65.6% (23.0% on average) of the peak. MHC-I multimer+ cells dominantly exhibit phenotypes of activated effector cells 1-2 weeks post-second vaccination and gradually acquire phenotypes of long-term memory cells, including stem cell-like memory T (TSCM) cells. Importantly, the frequency of TSCM cells 1-2 weeks post-second vaccination significantly correlates with the 6-month durability of CD8+ T cells, indicating that early generation of TSCM cells determines the longevity of vaccine-induced memory CD8+ T cell responses.

Lenalidomide bypasses CD28 co-stimulation to reinstate PD-1 immunotherapy by activating Notch signaling.

Programmed cell death protein 1 (PD-1) checkpoint blockade therapy requires the CD28 co-stimulatory receptor for CD8+ T cell expansion and cytotoxicity. However, CD28 expression is frequently lost in exhausted T cells and during immune senescence, limiting the clinical benefits of PD-1 immunotherapy in individuals with cancer. Here, using a cereblon knockin mouse model that regains in vivo T cell response to lenalidomide, an immunomodulatory imide drug, we show that lenalidomide reinstates the anti-tumor activity of CD28-deficient CD8+ T cells after PD-1 blockade. Lenalidomide redirects the CRL4Crbn ubiquitin ligase to degrade Ikzf1 and Ikzf3 in T cells and unleashes paracrine interleukin-2 (IL-2) and intracellular Notch signaling, which collectively bypass the CD28 requirement for activation of intratumoral CD8+ T cells and inhibition of tumor growth by PD-1 blockade. Our results suggest that PD-1 immunotherapy can benefit from a lenalidomide combination when treating solid tumors infiltrated with abundant CD28- T cells.

IL-17A-producing sinonasal MAIT cells in patients with chronic rhinosinusitis with nasal polyps.

BACKGROUND: Diverse immune cells contribute to the pathogenesis of chronic rhinosinusitis (CRS), an inflammatory disease of the nasal cavity and paranasal sinuses. However, whether mucosal-associated invariant T (MAIT) cells are present in human sinonasal tissues remains unclear. Furthermore, the characteristics of sinonasal MAIT cells have not been studied in patients with CRS. OBJECTIVE: We investigated the phenotype, function, and clinical implications of MAIT cells in patients with CRS. METHODS: Peripheral blood and sinonasal tissue were obtained from patients with CRS with (CRSwNP) or without nasal polyps (CRSsNP) and healthy controls. MAIT cells were analyzed by flow cytometry. RESULTS: We found that MAIT cells are present in human sinonasal tissues from healthy controls and patients with CRS. The sinonasal MAIT cell population, but not peripheral blood MAIT cells, from patients with CRSsNP, noneosinophilic CRSwNP (NE-NP), or eosinophilic CRSwNP (E-NP) had a significantly higher frequency of activated cells marked by CD38 expression. In functional analysis, the sinonasal MAIT cell population from NE-NP and E-NP had a significantly higher frequency of IL-17A+ cells but lower frequency of IFN-γ+ or TNF+ cells than control sinonasal tissues. Furthermore, CD38 expression and IL-17A production by sinonasal MAIT cells significantly correlated with disease extent evaluated by the Lund-Mackay computed tomography score in patients with E-NP. CONCLUSIONS: Sinonasal MAIT cells exhibit an activated phenotype and produce higher levels of IL-17A in patients with CRSwNP. These alterations are associated with the extent of disease in patients with E-NP.

SARS-CoV-2-specific T cell memory is sustained in COVID-19 convalescent patients for 10 months with successful development of stem cell-like memory T cells.

Memory T cells contribute to rapid viral clearance during re-infection, but the longevity and differentiation of SARS-CoV-2-specific memory T cells remain unclear. Here we conduct ex vivo assays to evaluate SARS-CoV-2-specific CD4+ and CD8+ T cell responses in COVID-19 convalescent patients up to 317 days post-symptom onset (DPSO), and find that memory T cell responses are maintained during the study period regardless of the severity of COVID-19. In particular, we observe sustained polyfunctionality and proliferation capacity of SARS-CoV-2-specific T cells. Among SARS-CoV-2-specific CD4+ and CD8+ T cells detected by activation-induced markers, the proportion of stem cell-like memory T (TSCM) cells is increased, peaking at approximately 120 DPSO. Development of TSCM cells is confirmed by SARS-CoV-2-specific MHC-I multimer staining. Considering the self-renewal capacity and multipotency of TSCM cells, our data suggest that SARS-CoV-2-specific T cells are long-lasting after recovery from COVID-19, thus support the feasibility of effective vaccination programs as a measure for COVID-19 control.

Effects of transarterial chemoembolization on regulatory T cell and its subpopulations in patients with hepatocellular carcinoma.

BACKGROUND: Regulatory T cell (Treg) plays an essential role in regulating anti-tumor immunity. The aim of this study was to investigate the effect of transarterial chemoembolization (TACE) on Treg in hepatocellular carcinoma (HCC) patients. METHOD: The frequency of peripheral blood Tregs in 27 HCC patients who underwent TACE were measured at baseline and 1 month after TACE. The frequency of peripheral blood Tregs at baseline were compared with those in 23 healthy controls. Tregs were further classified into three subpopulations [Treg (I), Treg (II), Treg (III)] based on expression levels or markers and their function. The patients were divided into two groups according to tumor response after TACE; complete response group and incomplete response group. The correlations between the frequency of Treg and clinical factors were analyzed. RESULTS: The frequency of Treg in HCC patients (7.52%) was significantly higher than in healthy controls (4.99%) at baseline. Regarding Treg subpopulations, the frequency of Treg (II) was significantly higher in HCC patients (2.51%) than in healthy controls (0.60%). In comparison of Treg numbers at baseline and post-TACE by tumor response, the change of Treg (III) in complete response group from baseline to post-TACE was significantly decreased (63.8 → 53.2/mm3). Patients with a high post-TACE Treg (III) (3.8 months) exhibited a significantly shorter median time to progression than those with a low post-TACE Treg (III) (11.6 months). In multivariate analyses, hypoalbuminemia (hazard ratio 3.324; 95% CI 1.098-10.063, p = 0.034) and high post-TACE Treg (III) (hazard ratio 3.080; 95% CI 1.091-8.696, p = 0.034) were significant factors for associating with progression. CONCLUSIONS: The frequency of Tregs in HCC patients was significantly higher than in healthy controls. In addition, patients with a high post-TACE Treg (III) exhibited a significantly lower progression-free survival rate than those with a low post-TACE Treg (III).

Prominent Inflammatory Features of Monocytes/Macrophages in Acute Calcium Pyrophosphate Crystal Arthritis: a Comparison with Acute Gouty Arthritis.

Calcium pyrophosphate (CPP) crystals can present as acute inflammatory arthritis which is known as an acute CPP crystal arthritis. Although monocytes/macrophages have been shown to play a role in the initiation of crystal-mediated inflammatory responses, differences in their phenotypes between acute CPP crystal arthritis and acute gouty arthritis have not yet been investigated. We examined the immunological characteristics of synovial monocytes/macrophages in patients with acute CPP crystal and acute gouty arthritis. CD14+CD3-CD19-CD56- cell frequencies in synovial fluid mononuclear cells (SFMCs) were measured. Expression of pro- and anti-inflammatory cytokines and markers was determined. The SFMCs were dominated by a population of monocytes/macrophages in acute CPP crystal arthritis similar to that in acute gout. Synovial monocytes/macrophages showed the phenotypes of infiltrated monocytes as shown by expression of CD88, C-C chemokine receptor type 2, myeloid-related protein (MRP)8 and MRP14 but not proto-oncogene tyrosine-protein kinase MER. Comparatively, the CD14+ cells from patients with acute CPP crystal arthritis had similar high levels of IL-1ß and TNF-α production but significantly lower expression of IL-10 and M2 marker (CD163). The monocytes/macrophages had the capacity to produce IL-8 in response to CPP crystals. Proinflammatory features were more dominant in monocytes/macrophages during acute CPP crystal arthritis than those during acute gouty arthritis.

Vertical alveolar ridge augmentation using autogenous bone grafts and platelet-enriched fibrin glue with simultaneous implant placement.

OBJECTIVE: The aim of this study was to evaluate the combined use of autogenous bone and platelet-enriched fibrin glue as grafting material for vertical alveolar ridge augmentation with simultaneous implant placement in a canine alveolar ridge defect model. STUDY DESIGN: In 6 mongrel dogs, bilateral vertical alveolar ridge defects were created in the mandible. After 3 months of healing, 2 dental implants were placed in each defect of the mandible, creating 6-mm supra-alveolar peri-implant defects. The 2 implants per defect were subjected to surgical treatments involving either a combination of autogenous bone grafts and platelet-enriched fibrin glue, or a conventional flap procedure only (control). After a healing period of 6 months, the dogs were humanely killed for histological and histometric analyses. RESULTS: Implant placement alone produced limited vertical alveolar height (0.6 +/- 0.4 mm). However, alveolar augmentation including a combination of autogenous bone grafts and platelet-enriched fibrin glue with simultaneous implant placement resulted in alveolar ridge augmentation amounting to 4.2 +/- 1.0 mm, comprising 63% of the defect height. New bone-implant contact was 40.5% in the defects treated with combined autogenous bone grafts and platelet-enriched fibrin glue, and was 48.4% in the resident bone; this difference was not statistically significant. CONCLUSION: The present study demonstrates that vertical alveolar ridge augmentation using autogenous bone grafts and platelet-enriched fibrin glue with simultaneous implant placement might effectively increase vertical alveolar ridge height and allow for an acceptable level of osseointegration.

Platelet-enriched fibrin glue and platelet-rich plasma in the repair of bone defects adjacent to titanium dental implants.

PURPOSE: The aim of this study was to compare the effects of platelet-enriched fibrin glue and platelet-rich plasma (PRP) on the repair of bone defects adjacent to titanium dental implants. MATERIALS AND METHODS: In 6 mongrel dogs, 3 screw-shaped titanium dental implants per dog were placed into the osteotomy sites in the tibia. Before implantation, a standardized gap (2.0 mm) was created between the implant surface and the surrounding bone walls. Six gaps were left empty (control group), 6 gaps were filled with autogenous particulate bone mixed with PRP (PRP group), and 6 gaps were filled with autogenous particulate bone mixed with platelet-enriched fibrin glue (fibrin glue group). RESULTS: After 6 weeks, the bone-implant contact was 59.7% in the fibrin glue group, 29.2% in the PRP group, and 10.2% in the control defects; this difference was statistically significant (P < .05). DISCUSSION AND CONCLUSION: Greater bone-implant contact was achieved with platelet-enriched fibrin glue than with PRP. The results indicate that platelet-enriched fibrin glue can induce a stronger peri-implant bone reaction than PRP in the treatment of bone defects adjacent to titanium dental implants.

Flapless implant surgery: an experimental study.

OBJECTIVE: The purpose of this study was to examine the effect of flapless implant surgery on crestal bone loss and osseointegration in a canine mandible model. STUDY DESIGN: In 6 mongrel dogs, bilateral, edentulated, flat alveolar ridges were created in the mandible. After 3 months of healing, 2 implants in each side were placed by either flap or flapless procedures. After a healing period of 8 weeks, microcomputerized tomography at the implantation site was performed. Osseointegration was calculated as percentage of implant surface in contact with bone. Additionally, bone height was measured in the peri-implant bone. RESULTS: The mean osseointegration was greater at flapless sites (70.4%) than at sites with flaps (59.5%) (P < .05). The mean peri-implant bone height was greater at flapless sites (10.1 mm) than at sites with flaps (9.0 mm) (P < .05). CONCLUSION: Flapless surgery can achieve results superior to surgery with reflected flaps. The specific improvements of this technique include enhanced osseointegration of dental implants and increased bone height.

The effect of platelet-rich plasma on bone healing around implants placed in bone defects treated with Bio-Oss: a pilot study in the dog tibia.

OBJECTIVE: The aim of this study was to examine the influence of platelet-rich plasma (PRP) used as an adjunct to Bio-Oss for the repair of bone defects adjacent to titanium dental implants. STUDY DESIGN: In 6 mongrel dogs, 12 screw-shaped titanium dental implants were inserted into the osteotomy sites in the dogs' tibias. Before implantation, a standardized gap (2.0 mm) was created between the implant surface and the surrounding bony walls. The gaps were filled with either Bio-Oss cancellous granules alone or Bio-Oss cancellous granules mixed with PRP. RESULTS: After 4 months, the Bio-Oss-treated defects revealed a significantly higher percentage of bone-implant contact than the defects treated with Bio-Oss and PRP (60.1% vs. 30.8%; P < .05). CONCLUSION: The results indicate that when PRP is used as an adjunct to Bio-Oss in the repair of bone defects adjacent to titanium dental implants, PRP may decrease periimplant bone healing.

Maxillary sinus floor augmentation using autogenous bone grafts and platelet-enriched fibrin glue with simultaneous implant placement.

OBJECTIVE: The aim of this study was to evaluate the use of autogenous bone in combination with platelet-enriched fibrin glue as a grafting material for maxillary sinus augmentation with simultaneous implant placement in dogs. STUDY DESIGN: The mucous membranes of 12 sinuses in 6 dogs were elevated bilaterally. In the right sinus, autogenous bone mixed with platelet-enriched fibrin glue was grafted into the space between the membrane and the sinus wall. In the left sinus, autogenous bone alone was grafted as a control. At the same time, 2 dental implants were inserted into the grafting material through the maxillary sinus floor. The animals were killed 6 months after surgery. RESULTS: The mean bone-implant contact was 40.5% on the fibrin glue side and 32.3% on the control side (P < .05). The mean height of newly formed bone in the augmented area was 12.2 mm on the fibrin glue side and 10.7 mm on the control side (P < .05). CONCLUSION: The results indicate that the use of autogenous bone mixed with platelet-enriched fibrin glue can achieve results superior to those for grafts of autogenous bone alone. The specific improvements of this technique include enhanced osseointegration of dental implants and increased height of new bone.

A retrospective study of the effects on sinus complications of exposing dental implants to the maxillary sinus cavity.

OBJECTIVE: The aim of this study was to investigate whether dental implant exposure to the maxillary sinus cavity increased the risk of maxillary sinus complications. STUDY DESIGN: Nine patients with 23 implants that had been inserted into the maxillary sinus more than 4 mm without lifting the sinus mucous membranes were evaluated for sinus complications 6 to 10 months after implant insertion, using a questionnaire and computerized tomography (CT). RESULTS: There were no clinical signs of sinusitis in any patient. However, CT scans showed postoperative sinus mucous thickening around 14 of the 23 implants. CONCLUSION: This study showed that implant exposure to the maxillary sinus cavity can cause sinus mucous thickening around the implants.

Treatment of experimental peri-implantitis using autogenous bone grafts and platelet-enriched fibrin glue in dogs.

OBJECTIVE: The purpose of this study was to evaluate the effects of autogenous bone grafts and platelet-enriched fibrin glue in the treatment of peri-implantitis. STUDY DESIGN: Thirty-six screw-type commercially pure titanium implants with rough acid-etched surfaces were inserted into 6 mongrel dogs 3 months after extraction of mandibular premolars. After 3 months of healing, peri-implantitis was induced by placing gauze and wire around the implants. Once peri-implantitis was created, surgical treatments involving a combination of autogenous bone grafts and platelet-enriched fibrin glue, autogenous bone grafts alone, or a conventional flap procedure only (control) were carried out. Six months later, biopsies of the implant sites were taken and prepared for ground sectioning and analysis. RESULTS: The amount of reosseointegration was significantly higher in peri-implantitis defects treated with combined autogenous bone grafts and platelet-enriched fibrin glue as compared with the other 2 treatment procedures. A mean bone-to-implant contact of 50.1% was obtained in the peri-implantitis lesions treated with combined autogenous bone grafts and platelet-enriched fibrin glue. The corresponding values for the autogenous bone grafts and control groups were 19.3% and 6.5%, respectively. CONCLUSION: The present study demonstrates that surgical treatment involving the combined use of autogenous bone grafts and platelet-enriched fibrin glue might effectively promote reosseointegration in lesions resulting from peri-implantitis.

Cyanoacrylate adhesive for closing sinus membrane perforations during sinus lifts.

INTRODUCTION: Sinus lift procedures depend greatly on fragile structures and anatomical variations. The procedure may cause sinus membrane perforations, which can lead to graft infection and early failure. AIM: To assess the efficacy of cyanoacrylate adhesive in the management of large perforations of the maxillary sinus membrane during sinus lifts. MATERIAL AND METHODS: Six rabbits were used in the study. Sinus membrane perforations (about 1.5 cm in length) were repaired with cyanoacrylate adhesive on one side of the maxillary sinus. On the contralateral side, an identical laceration was not repaired. Histological evaluation was performed 2 weeks after the operation. RESULTS: Wounds repaired with cyanoacrylate adhesive showed newly formed continuous epithelium across the previous perforation site and there was sinusitis on the contralateral side. CONCLUSION: These results support the clinical use of cyanoacrylate adhesive for repairing sinus membrane perforations.

The effects of exposing dental implants to the maxillary sinus cavity on sinus complications.

OBJECTIVE: The aim of this study was to investigate whether dental implant exposure to the maxillary sinus cavity increases the risk of maxillary sinus complications. STUDY DESIGN: An implant was placed bilaterally in the maxillary sinus of 8 adult female mongrel dogs in a way that it penetrated the bone and mucous membrane of the maxillary sinus floor to the extent of 2 mm, 4 mm, or 8 mm. The implants were left in place for 6 months. RESULTS: Radiographic and histologic examinations did not show any signs of pathologic findings in the maxillary sinus of the 8 dogs. CONCLUSION: This study indicates that implant protrusion into the maxillary sinus cavity is not related to the development of sinus complications in canines.