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Αλέξανδρος Γ. Σφακιανάκης

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Sunday, October 16, 2022

High titers of neutralizing antibodies in the blood fail to eliminate SARS‐CoV‐2 viral RNA in the upper respiratory tract

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Abstract

Retest positive SARS-CoV-2 viral RNA, as a unique phenomenon among the discharged individuals, has been demonstrated to be safe in the community. Still, the underlying mechanism of viral lingering is less investigated. In this study, first, we find that the frequency of viral RNA positive retest differs among variants. Higher ratios of viral RNA positive retest were more frequently observed among Delta (61.41%, 514 of 837 cases) and Omicrons (39.53%, 119 of 301 cases) infection than ancestral viral infection (7.27%, 21 of 289 cases). Second, the tissues where viral RNA reoccurred were altered. Delta RNA reoccurred mainly in the upper respiratory tract (90%), but ancestral virus RNA mainly in the gastrointestinal tract (71%). Third, vaccination did not reduce the frequency of viral RNA-positive retests, despite high concentrations of viral-specific antibodies in the blood. Finally, 37 of 55 (67.27%) Delta-infected patients receiving neutralizing antibody (nAb ) therapy become viral RNA retest positive when high concentrations of neutralizing antibodies still patrol in the blood. Altogether, our findings suggest that the presentence of high titers of neutralizing antibodies in the blood is incompetent in clearing residual viral RNA in the upper respiratory tract.

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Persistence of Neutrophil extracellular traps and anti‐cardiolipin auto‐antibodies in post‐acute phase COVID‐19 patients

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Abstract

This exploratory prospective study based on 279 individuals showed that plasma levels of neutrophil elastase, myeloperoxidase and circulating DNA of nuclear and mitochondrial origins in non-severe (NS), severe (S) and post-acute phase (PAP) COVID-19 patients were statistically different as compared to the levels in healthy individuals, and revealed the high diagnostic power of these markers in respect to the disease severity. The diagnostic power of NE, MPO, and cir-nDNA as determined by the Area Under Receiver Operating Curves (AUROC) was 0.95, 097 and 0.64; 0.99, 1.0 and 0.82; and 0.94, 1.0, and 0.93, in NS, S and PAP patient subgroups, respectively. In addition, a significant fraction of NS, S as well as of PAP patients exhibited aCL IgM/IgG and anti-B2GP IgM/IgG positivity. We first demonstrate persistence of these NETs (Neutrophil extracellular traps) markers in PAP patients and consequently of sustained innate immune response imbalance, and a prolonged low-level pro-thrombotic potential activity highlighting the need to monitor these markers in all COVID-19 PAP individuals, to investigate post-acute COVID-19 pathogenesis following intensive care, and to better identify which medical resources will ensure complete patient recovery.

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Bioactivity analysis of bioceramic sealers and repair cements on the phenotype and cytokine secretion profile of CD14+ monocytes: An ex‐vivo study

alexandrossfakianakis shared this article with you from Inoreader

Abstract

Aim

This study evaluated the immune bioactivity of testing media (TM) obtained from different calcium silicate-based sealers and cements on monocyte morphology, activation, differentiation, and cytokine secretion.

Methods

Blood-derived CD14+ monocytes were isolated and cultured for 5 days with 25% TM from the following calcium silicate-based materials: TotalFill BC RRM Fast-Set Putty, Biodentine, TotalFill BC Sealer, and BioRoot-RCS. A resin-based endodontic cement was used as a control. The expression of surface markers such as CD86, HLA-DR, CD16, CD309, and CD209, and cytokine secretion were analysed by flow cytometry. Data were analysed using the one-way repeated measures (RM) analysis of variance (ANOVA) multiple comparison test and a Holm-Sidak multiple comparison post-hoc test (p<0.05).

Results

This comparative analysis revealed that monocytes co-cultured with calcium silicate-based materials showed a spindle-shaped morphology compared to the round shape observed in the control. Regarding activation markers, BioRoot-RCS and Biodentine significantly increased CD86 expression compared with the control sample, whereas no significant differences (p>0.05) were observed in HLA-DR expression. In addition, no differences were observed among the differentiation markers. When the inflammatory cytokines were analysed, BioRoot-RCS increased the secretion of IL-1β, IL-6, IL-10, and TNF-α, whereas BioRoot-RCS and Biodentine significantly decreased IL-8 production (p<0.05).

Conclusions

These data showed that the calcium silicate-based materials tested changed the morphology of CD14+ monocytes; however, only BioRoot-RCS and Biodentine significantly upregulated CD86. In addition, BioRoot-RCS was the sealer with the highest immunomodulatory properties for cytokine production which means that it can contribute with the in vivo healing process and regeneration of periapical lesions.
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Characteristics and clinical significance of plasma IL‐18, sCD14 and sCD163 levels in patients with HIV‐1 infection

alexandrossfakianakis shared this article with you from Inoreader

Abstract

Background and aim

Biomarkers of monocyte-macrophages activation and inflammation in plasma such as interleukin-18 (IL-18), soluble leukocyte differentiation antigen 14 (sCD14) and sCD163 are associated with disease severity and prognosis in HIV-1 infected patients, however, their relationships with efficacy of antiretroviral therapy (ART) need further investigation. We aimed to characterize and explore the clinical significance of plasma IL-18, sCD14 and sCD163 in this population.

Methods

This was a retrospective cohort study consisting of HIV-1 infected patients enrolled in a randomized, controlled, open-label, non-inferiority trial (ALTERLL study), with follow-up time points including initiation of ART (baseline), 12-, 24- and 48-weeks of treatment. Plasma levels of IL-18, sCD14 and sCD163 were measured using enzyme-linked immunosorbent assay method. Viral suppression was defined as HIV-1 RNA <20 copies/mL.

Results

Among the 193 studied pa tients (median age of 29.0 years, 180 males), IL-18 and sCD163 had U-shaped regression curves and sCD14 had an inverted U-shaped regression curve while virus was decreasing and immune function recovered. Patients with higher levels of IL-18 or lower levels of sCD163 at baseline were less likely to achieve viral suppression at week 12 or week 24 of treatment, respectively. In multivariate analysis, baseline sCD163 ≤500 pg/mL (aOR 0.33, 95%CI 0.16-0.68) was independently associated with lower rate of viral suppression at week 24 of treatment.

Conclusion

We demonstrated different dynamic changes among IL-18, sCD14 and sCD163 after ART. Baseline sCD163 level could be a potential predictor of early virological response to ART. Further validation and mechanistic research are needed.

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Characterization and application of a series of monoclonal antibodies against SARS‐CoV‐2 nucleocapsid protein

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Abstract

The ongoing coronavirus disease 2019 (COVID-19) pandemic has a significant global social and economic impact, and the emergence of new and more destructive mutant strains highlights the need for accurate virus detection. Here, 90 monoclonal antibodies (MAbs) that exclusively reacted with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein (NP) were generated. These MAbs did not cross-react with NPs of common human coronaviruses (HCoV, i.e., 229E, OC43, HKU1, and NL63) and Middle East Respiratory Syndrome Coronavirus. Subsequently, overlapped peptides in individual fragments (N1–N4) of NP were synthesized. N1-3 (25-GSNQNGERSGARSKQ-39), N3-1 (217-AALALLLLDRLNQL-230), and N4-8 (393-TLLPAADLDDFSKQL-407) were identified as major epitopes using enzyme-linked immunoassay (ELISA) and recognized by 47, 1, and 18 MAbs, respectively. The 24 remaining MAbs exhibited no reactivity with all synthetic peptides. Among MAb-epitope pairs, only MAbs targeting epitope N1-3 displayed no cross-reaction with NPs of SARS-CoV-1 and other SARS-related CoVs. All omicron variants contained a three-amino acid deletion (31ERS33) in the N1-3 region. Thus, MAbs targeting N1-3 failed to recognize these variants. Furthermore, a double-antibody sandwich ELISA for antigen detection was established using the optimal MAbs. Overall, a series of MAbs targeting SARS-CoV-2 NP was prepared, characterized with epitope mapping, and applied for the detection of SARS-CoV-2 antigens, and some novel B-cell epitopes of the viral NP were identified.

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Development and validation of a nomogram for the prediction of lymph node metastasis within 2‐year postoperatively in cT1‐T2N0 oral squamous cell carcinoma

alexandrossfakianakis shared this article with you from Inoreader

Abstract

Background

The current neck management for early oral squamous cell carcinoma (OSCC) has always been a controversial issue. A comprehensive model is necessary for predicting an individual's metastasis risk and appropriate patient counseling.

Methods

A nomogram for predicting 2-year LNM in patients with cT1-2N0 OSCC was developed and validated using clinicopathological data from 642 patients from 2000 to 2018 in four hospitals, China.

Results

Three variables (pathology grade, depth of invasion, tumor-infiltrating lymphocytes) were included in nomogram. C-indices were 0.826 (95% CI: 0.786–0.866) and 0.726 (95% CI: 0.653–0.780) in the internal and external validation. Kaplan–Meier method found the 2-year LNM rate of high-risk group (35.8%) was much higher than that of the low-risk group (14.5%). The nomogram model has an advantage over the 8th AJCC TNM stage in predicting the individual 2-year LNM probability for early OSCC.

Conclusion

Patients with low-risk nomogram score may receive neck observation; those with high-risk score should receive END.
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USP36 promotes tumorigenesis and drug sensitivity of glioblastoma by deubiquitinating and stabilizing ALKBH5

alexandrossfakianakis shared this article with you from Inoreader
Abstract
Background
ALKBH5 is aberrantly activated and exerts critical roles in facilitating the development of glioblastoma. However, the underlying activation mechanism by which ALKBH5 protein is increased in glioblastoma is not completely understood. Our study aimed to elucidate the signaling pathways involved in mediating ALKBH5 protein stability.
Methods
The contribution of deubiquitinating enzymes (DUB) to the fluctuation of ALKBH5 protein expression were globally profiled with western blot analysis. Mass spectrometry and immunoprecipitation were performed to identify the USP36 and ALKBH5 interaction. The effects of USP36 on the stability of ALKBH5 were detected with in vivo and in vitro ubiquitination assays. Cell proliferation assays, neurosphere formation, limited dilution assay, and intracranial tumor growth assays were implemented to assess the co llaborative capacities of USP36 and ALKBH5 in tumorigenesis.
Results
Ubiquitin-specific peptidase 36 (USP36), as a potential ALKBH5-activating DUB, played an essential role in stabilization of ALKBH5 and regulation of ALKBH5 mediated gene expression in glioblastoma. The depletion of USP36 drastically impaired cell proliferation, deteriorated the self-renewal of GSCs and sensitized GSCs to temozolomide (TMZ) treatment. Furthermore, the deletion of USP36 substantially decreased the in vivo tumor growth when monitored by bioluminescence imaging. Our findings indicate that USP36 regulates the protein degradation and expression of ALKBH5, and the USP36-ALKBH5 axis orchestrates glioma tumorigenesis.
Conclusion
Our findings identify USP36 as a DUB of ALKBH5 and its role in glioblastoma progression, which may serve as a potential therapeutic target for glioblastoma treatment.
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A Nationwide Evaluation of Bevacizumab-based Treatments in Paediatric Low-Grade Glioma in the UK: Safety. Efficacy, Visual Morbidity and Outcomes

alexandrossfakianakis shared this article with you from Inoreader
Abstract
Background
Bevacizumab is increasingly used in children with Paediatric Low-Grade Glioma (PLGG) despite limited evidence. A nationwide UK service evaluation was conducted to provide larger cohort 'real life' safety and efficacy data including functional visual outcomes.
Methods
Children receiving Bevacizumab-based treatments (BBT) for PLGG (2009-2020) from 11 centres were included. Standardised neuro-radiological (RANO-LGG) and visual (logMAR visual acuity) criteria were used to assess clinical-radiological correlation, survival outcomes and multivariate prognostic analysis.
Results
Eighty-eight children with PLGG received BBT either as 3 rd line with Irinotecan (85%) or alongside 1 st/2 nd line chemotherapies (15%). Toxicity was limited and minimal. Partial response (PR, 40%), stable disease (SD, 49%), and progressive disease (PD, 11%) were seen during BBT. However, 65% progressed at 8 months (median) from BBT cessation, leading to a radiology-based 3yr-progression-free survival (PFS) of 29%. Diencephalic syndrome (p= 0.03) was associated with adverse PFS. Pre-existing visual morbidity included unilateral (25%) or bilateral (11%) blindness. Improvement (29%) or stabilisation (49%) of visual acuity were achieved, more often in patients' best eyes. Vision deteriorated during BBT in 14 (22%), with 3-year visual-PFS of 53%; more often in patients' worst eyes. A superior visual outcome (p=0.023) was seen in Neurofibromatosis type 1-associated Optic Pathway Glioma (OPG). Concordance between visual and radiological responses was 36%; optimised to 48% using only best eye responses.
Conclusions
BBTs provide effective short-term PLGG control and delay further progression, with a better sustained visual (best >worst eye) than radiological response. Further research could optimise the role of BBTs towards a potentially sight-saving strategy in OPG.
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Triple Antibiotic Paste Composition and use in Root Canal Treatment

alexandrossfakianakis shared this article with you from Inoreader

Triple Antibiotic Paste

Triple Antibiotic Paste is a type of Intra Canal Medicament which is used in Endodontic Treatment or Root canal Treatment to kill the micro organisms in the periapical region of an infected Root Canal. Triple antibiotic Paste or (TAP) consists of three medicines - Metronidazole, Ciprofloxacin and Minocycline. This combinati on of medicines have been found to be the best regimen to eliminate root canal pathogens. The most common microorganisms found in infected Root Canals are Peptostreptococcus spp, Actinomyces, Staphylococcus Salivarius, S. Sanguis, P. Endodontalis. The micro organisms most commonly infecting Failed or Re-infected root canals are "Enterococcus Faecalis". Triple Antibiotic paste uses: It is used as an Intracanal Medicament and in Regenerative Endodontics (Pulp Capping). It has been found to be very effective against E Feacalis which is a very stubborn bacteria found in Re-infected root canals. Composition of Triple antibiotic Paste: TAP is a combination of three antibiotics - Minocycline + Ciprofloxacin + Metronidazole. A combination of three antibiotics is required as any Infection of the Root Canal consists of multiple microorganisms of different types and to treat all these bacteria any single Antibiotic is sometimes not enough. A Combination of Antibiotics usually helps in treating the diverse flora of the Root Canal.
Triple Antibiotic Paste

How is Triple Antibiotics Paste prepared:

Hoshino et al. first proposed the combination of TPA and mentioned that the three Antibiotics - Metronidazole (500 mg) 30% + Minocycline (100mg) 30% + Ciprofloxacin (200mg) 30% + Propylene Glycol paste 10% should be mixed in a ratio of 1:1:1. These three components should have a carrier Glycol and macrogol ointment in a ratio of 1:1. The combination was later modified by making changes in the Ratios by Takushige et al. who proposed that the ratios should be Metronidazole + Minocycline + Ciprofloxacin - 3:3:1. A new Intra Canal Medicament known as MTAD was found to be useful in eliminating Enterococcus Faecalis strains  in a study published by Newberry et al. The MTAD is a combination of 3% Doxycycline, Citric Acid and a detergent polysorbate 80. MTAD should be used as an irrigation fluid inside the canals. MTAD should be used as the final irrigation rinse after using 1.3% Naocl and Saline. This has been found to remove the smear layer without altering the structure of dentinal tubules.
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The Role of Fluorescent Angiography in Free Flap Reconstruction of the Head and Neck

alexandrossfakianakis shared this article with you from Inoreader
The Role of Fluorescent Angiography in Free Flap Reconstruction of the Head and Neck

A retrospective study of the use of fluorescent angiography in free flap reconstruction of the head and neck shows that although the use of this technology in every free tissue transfer is not justifiable, it can guide the clinical course in challenging scenarios.


Objectives

Highlight the use of fluorescent angiography in free flap reconstruction of the head and neck. Qualify how fluorescent angiography can be selectively added to management paradigms for head and neck free flap reconstruction.

Methods

Retrospective chart review of 993 free flaps completed from the time the SPY Elite® system first became available at our institution between September 2013, until August 2020. Cases that used the SPY Elite® system were grouped into three broad categories: evaluation during initial flap harvest while still attached to the donor site, evaluation after anastomosis in the head and neck area, and evaluation post-operatively for questionable flap viability.

Results

The SPY Elite® system was used in 64 cases. Forty flaps were evaluated intraoperatively during initial harvest and before anastomosis to the head and neck area. Of these, 20 had signs of poor perfusion of the entire skin paddle, 12 had large myogenous or skin flaps with questionable perfusion of the distal aspect, and 8 were evaluated for other reasons. In this group the use of SPY Elite® changed the management of the patient in 20 cases (50%). Ten flaps were evaluated intraoperatively after anastomosis to the head and neck to ascertain adequate flow to the entire flap. In this group management was changed in two (20%). Fourteen flaps were evaluated 3–5 days post operatively due to suspected failure of a component. In five cases (36%), the use of SPY Elite® determined management with either trimming or discarding the flap.

Conclusion

Assessment of flap perfusion via fluorescent angiography during initial flap harvest or when flap compromise is suspected post-operatively can guide decision making in free flap reconstruction of the head and neck and can be added to existing planning and management paradigms.

Level of Evidence

4 Laryngoscope, 2022
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