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Investigative Ophthalmology & Visual... Jul 2024The purpose of this study was to evaluate pupillary light reflex (PLR) to chromatic flashes in patients with early-onset high-myopia (eoHM) without (myopic controls =...
PURPOSE
The purpose of this study was to evaluate pupillary light reflex (PLR) to chromatic flashes in patients with early-onset high-myopia (eoHM) without (myopic controls = M-CTRL) and with (female-limited myopia-26 = MYP-26) genetic mutations in the ARR3 gene encoding the cone arrestin.
METHODS
Participants were 26 female subjects divided into 3 groups: emmetropic controls (E-CTRL, N = 12, mean age = 28.6 ± 7.8 years) and 2 myopic (M-CTRL, N = 7, mean age = 25.7 ± 11.5 years and MYP-26, N = 7, mean age = 28.3 ± 15.4 years) groups. In addition, one hemizygous carrier and one control male subject were examined. Direct PLRs were recorded after 10-minute dark adaptation. Stimuli were 1-second red (peak wavelength = 621 nm) and blue (peak wavelength = 470 nm) flashes at photopic luminance of 250 cd/m². A 2-minute interval between the flashes was introduced. Baseline pupil diameter (BPD), peak pupil constriction (PPC), and postillumination pupillary response (PIPR) were extracted from the PLR. Group comparisons were performed with ANOVAs.
RESULTS
Dark-adapted BPD was comparable among the groups, whereas PPC to the red light was slightly reduced in patients with myopia (P = 0.02). PIPR at 6 seconds elicited by the blue flash was significantly weaker (P < 0.01) in female patients with MYP-26, whereas it was normal in the M-CTRL group and the asymptomatic male carrier.
CONCLUSIONS
L/M-cone abnormalities due to ARR3 gene mutation is currently claimed to underlie the pathological eye growth in MYP-26. Our results suggest that malfunction of the melanopsin system of intrinsically photosensitive retinal ganglion cells (ipRGCs) is specific to patients with symptomatic MYP-26, and may therefore play an additional role in the pathological eye growth of MYP-26.
Topics: Humans; Female; Reflex, Pupillary; Rod Opsins; Adult; Young Adult; Dark Adaptation; Myopia; Male; Photic Stimulation; Adolescent; Arrestin; Mutation; Pupil; Light; Middle Aged; Myopia, Degenerative
PubMed: 38958970
DOI: 10.1167/iovs.65.8.6 -
Applied Biochemistry and Biotechnology Jul 2024Simultaneous targeting of several mutations can be useful in colorectal cancer (CRC) due to its heterogeneity and presence of somatic mutations. As CT26 mutations and...
Simultaneous targeting of several mutations can be useful in colorectal cancer (CRC) due to its heterogeneity and presence of somatic mutations. As CT26 mutations and expression profiles resemble those of human CRC, we focused on designing a polyepitope vaccine based on CT26 neoepitopes. Due to its low immunogenicity, outer membrane vesicles (rOMV) as an antigen delivery system and adjuvant was applied. Herein, based on previous experimental and our in silico studies four CT26 neoepitopes with the ability to bind MHC-I and MHC-II, TCR, and induce IFN-α production were selected. To increase their immunogenicity, the gp70 and PADRE epitopes were added. The order of the neoepitopes was determined through 3D structure analysis using ProSA, Verify 3D, ERRAT, and Ramachandran servers. The stable peptide-protein docking between the selected epitopes and MHC alleles strengthen our prediction. The CT26 polytope vaccine sequence was fused to the C-terminal of cytolysin A (ClyA) anchor protein and rOMVs were isolated from endotoxin-free ClearColi™ strain. The results of the C-ImmSim server showed that the ClyA-CT26 polytope vaccine could induce T and B cells immunity.The ClyA-CT26 polytope was characterized as a soluble, stable, immunogen, and non-allergen vaccine and optimized for expression in ClearColi™ 24 h after induction with 1 mM IPTG at 25 °C. Western blot analysis confirmed the expression of ClyA-CT26 polytope by ClearColi™ and also on ClearColi™-derived rOMVs. In conclusion, we found that ClearColi™-derived rOMVs with CT26 polytope can deliver CRC neoantigens and induce antitumor immunity, but in vivo immunological studies are needed to confirm vaccine efficacy.
PubMed: 38958886
DOI: 10.1007/s12010-024-04971-x -
Acta Crystallographica. Section F,... Jul 2024The third complementary-determining regions of the heavy-chain (CDR3H) variable regions (VH) of some cattle antibodies are highly extended, consisting of 48 or more...
The third complementary-determining regions of the heavy-chain (CDR3H) variable regions (VH) of some cattle antibodies are highly extended, consisting of 48 or more residues. These `ultralong' CDR3Hs form β-ribbon stalks that protrude from the surface of the antibody with a disulfide cross-linked knob region at their apex that dominates antigen interactions over the other CDR loops. The structure of the Fab fragment of a naturally paired bovine ultralong antibody (D08), identified by single B-cell sequencing, has been determined to 1.6 Å resolution. By swapping the D08 native light chain with that of an unrelated antigen-unknown ultralong antibody, it is shown that interactions between the CDR3s of the variable domains potentially affect the fine positioning of the ultralong CDR3H; however, comparison with other crystallographic structures shows that crystalline packing is also a major contributor. It is concluded that, on balance, the exact positioning of ultralong CDR3H loops is most likely to be due to the constraints of crystal packing.
Topics: Animals; Cattle; Immunoglobulin Heavy Chains; Crystallography, X-Ray; Immunoglobulin Light Chains; Complementarity Determining Regions; Immunoglobulin Fab Fragments; Models, Molecular; Amino Acid Sequence; Protein Conformation
PubMed: 38958188
DOI: 10.1107/S2053230X2400606X -
Frontiers in Cellular and Infection... 2024Tuberculosis (TB) persists as a global health challenge, with its treatment hampered by the side effects of long-term combination drug therapies and the growing issue of...
BACKGROUND
Tuberculosis (TB) persists as a global health challenge, with its treatment hampered by the side effects of long-term combination drug therapies and the growing issue of drug resistance. Therefore, the development of novel therapeutic strategies is critical. This study focuses on the role of immune checkpoint molecules (ICs) and functions of CD8+ T cells in the search for new potential targets against TB.
METHODS
We conducted differential expression genes analysis and CD8+ T cell functional gene analysis on 92 TB samples and 61 healthy individual (HI) samples from TB database GSE83456, which contains data on 34,603 genes. The GSE54992 dataset was used to validated the findings. Additionally, a cluster analysis on single-cell data from primates infected with mycobacterium tuberculosis and those vaccinated with BCG was performed.
RESULTS
The overexpression of LAG-3 gene was found as a potentially important characteristic of both pulmonary TB (PTB) and extrapulmonary TB (EPTB). Further correlation analysis showed that LAG-3 gene was correlated with GZMB, perforin, IL-2 and IL-12. A significant temporal and spatial variation in LAG-3 expression was observed in T cells and macrophages during TB infection and after BCG vaccination.
CONCLUSION
LAG-3 was overexpressed in TB samples. Targeting LAG-3 may represent a potential therapeutic target for tuberculosis.
Topics: Lymphocyte Activation Gene 3 Protein; Humans; Mycobacterium tuberculosis; CD8-Positive T-Lymphocytes; Tuberculosis; Animals; Antigens, CD; BCG Vaccine; Macrophages; Interleukin-2; Gene Expression Profiling; Tuberculosis, Pulmonary; Interleukin-12; Perforin; Male
PubMed: 38957797
DOI: 10.3389/fcimb.2024.1410015 -
MAbs 2024Precise measurement of the binding activity changes of therapeutic antibodies is important to determine the potential critical quality attributes (CQAs) in...
Early determination of potential critical quality attributes of therapeutic antibodies in developability studies through surface plasmon resonance-based relative binding activity assessment.
Precise measurement of the binding activity changes of therapeutic antibodies is important to determine the potential critical quality attributes (CQAs) in developability assessment at the early stage of antibody development. Here, we report a surface plasmon resonance (SPR)-based relative binding activity method, which incorporates both binding affinity and binding response and allows us to determine relative binding activity of antibodies with high accuracy and precision. We applied the SPR-based relative binding activity method in multiple forced degradation studies of antibody developability assessment. The current developability assessment strategy provided comprehensive, precise characterization of antibody binding activity in the stability studies, enabling us to perform correlation analysis and establish the structure-function relationship between relative binding activity and quality attributes. The impact of a given quality attribute on binding activity could be confidently determined without isolating antibody variants. We identified several potential CQAs, including Asp isomerization, Asn deamidation, and fragmentation. Some potential CQAs affected binding affinity of antibody and resulted in a reduction of binding activity. Certain potential CQAs impaired antibody binding to antigen and led to a loss of binding activity. A few potential CQAs could influence both binding affinity and binding response and cause a substantial decrease in antibody binding activity. Specifically, we identified low abundance Asn33 deamidation in the light chain complementarity-determining region as a potential CQA, in which all the stressed antibody samples showed Asn33 deamidation abundances ranging from 4.2% to 27.5% and a mild binding affinity change from 1.76 nM to 2.16 nM.
Topics: Surface Plasmon Resonance; Humans; Antibodies, Monoclonal; Antibody Affinity; Protein Binding; Animals
PubMed: 38956880
DOI: 10.1080/19420862.2024.2374607 -
The Journal of Contemporary Dental... Apr 2024This study aims to assess the synergistic effect of utilizing a bioceramic sealer, NeoPutty, with photobiomodulation (PBM) on dental pulp stem cells (DPSCs) for...
AIMS
This study aims to assess the synergistic effect of utilizing a bioceramic sealer, NeoPutty, with photobiomodulation (PBM) on dental pulp stem cells (DPSCs) for odontogenesis.
MATERIALS AND METHODS
Dental pulp stem cells were collected from 10 premolars extracted from healthy individuals. Dental pulp stem cells were characterized using an inverted-phase microscope to detect cell shape and flow cytometry to detect stem cell-specific surface antigens. Three experimental groups were examined: the NP group, the PBM group, and the combined NP and PBM group. A 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) experiment was conducted to assess the viability of DPSCs. The odontogenic differentiation potential was analyzed using Alizarin red staining, RT-qPCR analysis of odontogenic genes DMP-1, DSPP, and alkaline phosphatase (ALP), and western blot analysis for detecting BMP-2 and RUNX-2 protein expression. An analysis of variance (ANOVA) followed by a -test was employed to examine and compare the mean values of the results.
RESULTS
The study showed a notable rise in cell viability when NP and PBM were used together. Odontogenic gene expression and the protein expression of BMP-2 and RUNX-2 were notably increased in the combined group. The combined effect of NeoPutty and PBM was significant in enhancing the odontogenic differentiation capability of DPSCs.
CONCLUSION
The synergistic effect of NeoPutty and PBM produced the most positive effect on the cytocompatibility and odontogenic differentiation potential of DPSCs.
CLINICAL SIGNIFICANCE
Creating innovative regenerative treatments to efficiently and durably repair injured dental tissues. How to cite this article: Alshawkani HA, Mansy M, Al Ankily M, . Regenerative Potential of Dental Pulp Stem Cells in Response to a Bioceramic Dental Sealer and Photobiomodulation: An Study. J Contemp Dent Pract 2024;25(4):313-319.
Topics: Dental Pulp; Humans; Stem Cells; Low-Level Light Therapy; Cell Differentiation; Bone Morphogenetic Protein 2; Odontogenesis; Root Canal Filling Materials; Alkaline Phosphatase; In Vitro Techniques; Cell Survival; Regeneration; Ceramics; Extracellular Matrix Proteins; Cells, Cultured; Core Binding Factor Alpha 1 Subunit; Sialoglycoproteins; Phosphoproteins
PubMed: 38956844
DOI: 10.5005/jp-journals-10024-3676 -
Stem Cell Research & Therapy Jul 2024The human induced pluripotent stem cells (hiPSCs) can generate all the cells composing the human body, theoretically. Therefore, hiPSCs are thought to be a candidate...
BACKGROUND
The human induced pluripotent stem cells (hiPSCs) can generate all the cells composing the human body, theoretically. Therefore, hiPSCs are thought to be a candidate source of stem cells for regenerative medicine. The major challenge of allogeneic hiPSC-derived cell products is their immunogenicity. The hypoimmunogenic cell strategy is allogenic cell therapy without using immune suppressants. Advances in gene engineering technology now permit the generation of hypoimmunogenic cells to avoid allogeneic immune rejection. In this study, we generated a hypoimmunogenic hiPSC (HyPSC) clone that had diminished expression of human leukocyte antigen (HLA) class Ia and class II and expressed immune checkpoint molecules and a safety switch.
METHODS
First, we generated HLA class Ia and class II double knockout (HLA class Ia/II DKO) hiPSCs. Then, a HyPSC clone was generated by introducing exogenous β-2-microglobulin (B2M), HLA-G, PD-L1, and PD-L2 genes, and the Rapamycin-activated Caspase 9 (RapaCasp9)-based suicide gene as a safety switch into the HLA class Ia/II DKO hiPSCs. The characteristics and immunogenicity of the HyPSCs and their derivatives were analyzed.
RESULTS
We found that the expression of HLA-G on the cell surface can be enhanced by introducing the exogenous HLA-G gene along with B2M gene into HLA class Ia/II DKO hiPSCs. The HyPSCs retained a normal karyotype and had the characteristics of pluripotent stem cells. Moreover, the HyPSCs could differentiate into cells of all three germ layer lineages including CD45 hematopoietic progenitor cells (HPCs), functional endothelial cells, and hepatocytes. The HyPSCs-derived HPCs exhibited the ability to evade innate and adaptive immunity. Further, we demonstrated that RapaCasp9 could be used as a safety switch in vitro and in vivo.
CONCLUSION
The HLA class Ia/II DKO hiPSCs armed with HLA-G, PD-L1, PD-L2, and RapaCasp9 molecules are a potential source of stem cells for allogeneic transplantation.
Topics: Humans; Induced Pluripotent Stem Cells; B7-H1 Antigen; Adaptive Immunity; Immunity, Innate; HLA-G Antigens; Programmed Cell Death 1 Ligand 2 Protein; Animals; Mice
PubMed: 38956724
DOI: 10.1186/s13287-024-03810-4 -
European Journal of Medical Research Jul 2024Breast cancer (BC) has a high mortality rate and is one of the most common malignancies in the world. Initially, BC was considered non-immunogenic, but a paradigm shift... (Review)
Review
Breast cancer (BC) has a high mortality rate and is one of the most common malignancies in the world. Initially, BC was considered non-immunogenic, but a paradigm shift occurred with the discovery of tumor-infiltrating lymphocytes (TILs) and regulatory T cells (Tregs) in the BC tumor microenvironment. CTLA-4 (Cytotoxic T-lymphocyte-associated protein 4) immunotherapy has emerged as a treatment option for BC, but it has limitations, including suboptimal antitumor effects and toxicity. Research has demonstrated that anti-CTLA-4 combination therapies, such as Treg depletion, cancer vaccines, and modulation of the gut microbiome, are significantly more effective than CTLA-4 monoclonal antibody (mAB) monotherapy. Second-generation CTLA-4 antibodies are currently being developed to mitigate immune-related adverse events (irAEs) and augment antitumor efficacy. This review examines anti-CTLA-4 mAB in BC, both as monotherapy and in combination with other treatments, and sheds light on ongoing clinical trials, novel CTLA-4 therapeutic strategies, and potential utility of biomarkers in BC.
Topics: Humans; CTLA-4 Antigen; Breast Neoplasms; Female; Immunotherapy; Tumor Microenvironment; Antibodies, Monoclonal; T-Lymphocytes, Regulatory; Lymphocytes, Tumor-Infiltrating
PubMed: 38956700
DOI: 10.1186/s40001-024-01901-9 -
Journal of Translational Medicine Jul 2024CD19-targeted chimeric antigen receptor T (CAR-T) cell therapy stands out as a revolutionary intervention, exhibiting remarkable remission rates in patients with...
BACKGROUND
CD19-targeted chimeric antigen receptor T (CAR-T) cell therapy stands out as a revolutionary intervention, exhibiting remarkable remission rates in patients with refractory/relapsed (R/R) B-cell malignancies. However, the potential side effects of therapy, particularly cytokine release syndrome (CRS) and infections, pose significant challenges due to their overlapping clinical features. Promptly distinguishing between CRS and infection post CD19 target CAR-T cell infusion (CTI) remains a clinical dilemma. Our study aimed to analyze the incidence of infections and identify key indicators for early infection detection in febrile patients within 30 days post-CTI for B-cell malignancies.
METHODS
In this retrospective cohort study, a cohort of 104 consecutive patients with R/R B-cell malignancies who underwent CAR-T therapy was reviewed. Clinical data including age, gender, CRS, ICANS, treatment history, infection incidence, and treatment responses were collected. Serum biomarkers procalcitonin (PCT), interleukin-6 (IL-6), and C-reactive protein (CRP) levels were analyzed using chemiluminescent assays. Statistical analyses employed Pearson's Chi-square test, t-test, Mann-Whitney U-test, Kaplan-Meier survival analysis, Cox proportional hazards regression model, Spearman rank correlation, and receiver operating characteristic (ROC) curve analysis to evaluate diagnostic accuracy and develop predictive models through multivariate logistic regression.
RESULTS
In this study, 38 patients (36.5%) experienced infections (30 bacterial, 5 fungal, and 3 viral) within the first 30 days of CAR T-cell infusion. In general, bacterial, fungal, and viral infections were detected at a median of 7, 8, and 9 days, respectively, after CAR T-cell infusion. Prior allogeneic hematopoietic cell transplantation (HCT) was an independent risk factor for infection (Hazard Ratio [HR]: 4.432 [1.262-15.565], P = 0.020). Furthermore, CRS was an independent risk factor for both infection ((HR: 2.903 [1.577-5.345], P < 0.001) and severe infection (9.040 [2.256-36.232], P < 0.001). Serum PCT, IL-6, and CRP were valuable in early infection prediction post-CAR-T therapy, particularly PCT with the highest area under the ROC curve (AUC) of 0.897. A diagnostic model incorporating PCT and CRP demonstrated an AUC of 0.903 with sensitivity and specificity above 83%. For severe infections, a model including CRS severity and PCT showed an exceptional AUC of 0.991 with perfect sensitivity and high specificity. Based on the aforementioned analysis, we proposed a workflow for the rapid identification of early infection during CAR-T cell therapy.
CONCLUSIONS
CRS and prior allogeneic HCT are independent infection risk factors post-CTI in febrile B-cell malignancy patients. Our identification of novel models using PCT and CRP for predicting infection, and PCT and CRS for predicting severe infection, offers potential to guide therapeutic decisions and enhance the efficacy of CAR-T cell therapy in the future.
Topics: Humans; Female; Male; Middle Aged; Immunotherapy, Adoptive; Adult; Antigens, CD19; Fever; Infections; Aged; ROC Curve; Young Adult; Retrospective Studies
PubMed: 38956649
DOI: 10.1186/s12967-024-05308-2 -
Journal of Nanobiotechnology Jul 2024Porcine reproductive and respiratory syndrome virus (PRRSV) is a prevalent swine pathogen, which has caused adverse impact on the global swine industry for almost 30...
BACKGROUND
Porcine reproductive and respiratory syndrome virus (PRRSV) is a prevalent swine pathogen, which has caused adverse impact on the global swine industry for almost 30 years. However, due to the immune suppression caused by the virus and the genetic diversity in PRRSV, no virus-targeting broad neutralizing strategy has been successfully developed yet. Antiviral peptide and nanobody have attracted extensive attention with the ease in production and the efficacy in practice. In this study, four new fusion proteins named nanobody peptide conjugates (NPCs) were developed by combining PRRSV specific non-neutralizing nanobodies with CD163-derived peptides targeting the receptor binding domain (RBD) of PRRSV proteins.
RESULTS
Four NPCs were successfully constructed using two nanobodies against PRRSV N and nsp9 individually, recombining with two antiviral peptides 4H7 or 8H2 from porcine CD163 respectively. All four NPCs demonstrated specific capability of binding to PRRSV and broad inhibitory effect against various lineages of PRRSV in a dose-dependent manner. NPCs interfere with the binding of the RBD of PRRSV proteins to CD163 in the PRRSV pre-attachment stage by CD163 epitope peptides in the assistance of Nb components. NPCs also suppress viral replication during the stage of post-attachment, and the inhibitory effects depend on the antiviral functions of Nb parts in NPCs, including the interference in long viral RNA synthesis, NF-κB and IFN-β activation. Moreover, an interaction was predicted between aa K31 and T32 sites of neutralizing domain 4H7 of NPC-N/nsp9-4H7 and the motif 171NLRLTG176 of PRRSV GP2a. The motif 28SSS30 of neutralizing domain 8H2 of NPC-N/nsp9-8H2 could also form hydrogens to bind with the motif 152NAFLP156 of PRRSV GP3. The study provides valuable insights into the structural characteristics and potential functional implications of the RBD of PRRSV proteins. Finally, as indicated in a mouse model, NPC intranasally inoculated in vivo for 12-24 h sustains the significant neutralizing activity against PRRSV. These findings inspire the potential of NPC as a preventive measure to reduce the transmission risk in the host population against respiratory infectious agents like PRRSV.
CONCLUSION
The aim of the current study was to develop a peptide based bioactive compound to neutralize various PRRSV strains. The new antiviral NPC (nanobody peptide conjugate) consists of a specific nanobody targeting the viral protein and a neutralizing CD163 epitope peptide for virus blocking and provides significant antiviral activity. The study will greatly promote the antiviral drug R&D against PRRSV and enlighten a new strategy against other viral diseases.
Topics: Porcine respiratory and reproductive syndrome virus; Animals; Single-Domain Antibodies; Swine; Antigens, Differentiation, Myelomonocytic; Receptors, Cell Surface; Antigens, CD; Antibodies, Neutralizing; Peptides; Porcine Reproductive and Respiratory Syndrome; Mice; Virus Replication; Cell Line
PubMed: 38956618
DOI: 10.1186/s12951-024-02662-7