scCURE identifies cell types responding to immunotherapy and enables outcome prediction.

A deep understanding of immunotherapy response/resistance mechanisms and a highly reliable therapy response prediction are vital for cancer treatment. Here, we developed scCURE (single-cell RNA sequencing [scRNA-seq] data-based Changed and Unchanged cell Recognition during immunotherapy). Based on Gaussian mixture modeling, Kullback-Leibler (KL) divergence, and mutual nearest-neighbors criteria, scCURE can faithfully discriminate between cells affected or unaffected by immunotherapy intervention. By conducting scCURE analyses in melanoma and breast cancer immunotherapy scRNA-seq data, we found that the baseline profiles of specific CD8+ T and macrophage cells (identified by scCURE) can determine the way in which tumor microenvironment immune cells respond to immunotherapy, e.g., antitumor immunity activation or de-activation; therefore, these cells could be predictive factors for treatment response. In this work, we demonstrated that the immunotherapy-associated cell-cell heterogeneities revealed by scCURE can be utilized to integrate the therapy response mechanism study and prediction model construction.

Differential genome-wide associated variants and enriched pathways of ECG parameters among people with versus without HIV.

OBJECTIVES: People with HIV (PWH) are more likely to develop ECG abnormalities. Substantial evidence exists for genetic contribution to ECG parameters among general population. However, whether and how would host genome associate with ECG parameters among PWH is unclear. Our research aims to analyze and compare genetic variants, mapped genes, and enriched pathways of ECG parameters among PWH and HIV-negative controls. DESIGN: A cross-sectional study. METHOD: We performed a large original genome-wide association study (GWAS) of ECG parameters among PWH ( n  = 1730) and HIV-negative controls ( n =  3746). Genome-wide interaction analyses were also conducted. RESULTS: A total of 18 novel variants were detected among PWH, six for PR interval including rs76345397 at ATL2 , 11 for QRS duration including rs10483994 at KCNK10 and rs2478830 at JCAD , and one for QTc interval (rs9815364). Among HIV-negative controls, we identified variants located at previously reported ECG-related genes ( SCN5A , CNOT1 ). Genetic variants had a significant interaction with HIV infection ( P  < 5 × 10 -8 ), implying that HIV infection and host genome might jointly influence ECG parameters. Mapped genes for PR interval and QRS duration among PWH were enriched in the biological process of viral genome replication and host response to virus, respectively, whereas enriched pathways for PR interval among HIV-negative controls were in the cellular component of voltage-gated sodium channel complex. CONCLUSION: The present GWAS indicated a distinctive impact of host genome on quantitative ECG parameters among PWH. Different from HIV-negative controls, host genome might influence the cardiac electrical activity by interfering with HIV viral infection, production, and latency among PWH.

Genome-wide associated variants of subclinical atherosclerosis among young people with HIV and gene-environment interactions.

BACKGROUND: Genome-wide association studies (GWAS) have identified some variants associated with subclinical atherosclerosis (SCA) in general population but lacking sufficient validation. Besides traditional risk factors, whether and how would genetic variants associate with SCA among people with HIV (PWH) remains to be elucidated. METHOD: A large original GWAS and gene-environment interaction analysis of SCA were conducted among Chinese PWH (n = 2850) and age/sex-matched HIV-negative controls (n = 5410). Subgroup analyses by age and functional annotations of variants were also performed. RESULTS: Different from HIV-negative counterparts, host genome had a greater impact on young PWH rather than the elders: one genome-wide significant variant (rs77741796, P = 2.20 × 10-9) and eight suggestively significant variants (P < 1 × 10-6) were identified to be specifically associated with SCA among PWH younger than 45 years. Seven genomic loci and 15 genes were mapped to play a potential role on SCA among young PWH, which were enriched in the biological processes of atrial cardiac muscle cell membrane repolarization and molecular function of protein kinase A subunit binding. Furthermore, genome-wide interaction analyses revealed significant HIV-gene interactions overall as well as gene-environment interactions with alcohol consumption, tobacco use and obesity among PWH. The identified gene-environment interaction on SCA among PWH might be useful for discovering high-risk individuals for the prevention of SCA, particularly among those with tobacco use and alcohol consumption. CONCLUSION: The present study provides new clues for the genetic contribution of SCA among young PWH and is the starting point of precision intervention targeting HIV-related atherosclerosis.

scCODE: an R package for data-specific differentially expressed gene detection on single-cell RNA-sequencing data.

Differential expression (DE) gene detection in single-cell ribonucleic acid (RNA)-sequencing (scRNA-seq) data is a key step to understand the biological question investigated. Filtering genes is suggested to improve the performance of DE methods, but the influence of filtering genes has not been demonstrated. Furthermore, the optimal methods for different scRNA-seq datasets are divergent, and different datasets should benefit from data-specific DE gene detection strategies. However, existing tools did not take gene filtering into consideration. There is a lack of metrics for evaluating the optimal method on experimental datasets. Based on two new metrics, we propose single-cell Consensus Optimization of Differentially Expressed gene detection, an R package to automatically optimize DE gene detection for each experimental scRNA-seq dataset.

[Comparison of the accuracy for evaluating cervical vertebral bone age and dental age of children in Shanghai].

PURPOSE: To compare the applicability and validity of dental age (DA) estimated by Willems method and cervical vertebral bone age (CVBA) evaluated by regression formula in estimating the chronological age of children in Shanghai. METHODS: Panoramic radiographs and lateral cephalograms were retrospectively collected from 320 subjects (160 males, 160 females), totaling 640 images. Discrepancies between chronological and estimated ages were statistically calculated by paired samples t test or Wilcoxon signed rank test using SPSS 25.0 software package. The accuracy of the two methods was comprehensively evaluated by comparing their standard deviation, mean absolute error (MAE) and the correct rate of acceptable range of estimated age error. RESULTS: The mean DA underestimated CA by 0.75±1.03 years for males and by 1.05±1.18 years for females; whereas the mean CVBA underestimated CA by 0.78±1.40 years for males and 0.53±1.31 years for females. MAE of Willems method was 1.15 years and the MAE of regression formula of CVBA was 1.20 years. The correct rate of clinically acceptable error of 0.5 years was 26.25% for Willems method and 27.19% for regression formula of CVBA. CONCLUSIONS: Willems method is more accurate than regression formula in indicating cervical vertebral skeletal age of adolescents in Shanghai children. Because of significant differences between CA and estimated ages, further modifications are urged to improve the accuracy of these two methods.

Multi-Mode Antibacterial Strategies Enabled by Gene-Transfection and Immunomodulatory Nanoparticles in 3D-Printed Scaffolds for Synergistic Exogenous and Endogenous Treatment of Infections.

As research on refractory Staphylococcus aureus-related implant infection intensifies, certain challenges remain, including low antibiotic concentrations within infected areas, immune escape achieved by intracellular bacteria, myeloid-derived suppressor cells (MDSCs) inducing regional immunosuppression, and recurrence of residual pathogenic bacteria after drug suspension. Herein, a novel antimicrobial system to simultaneously address these issues is proposed. Specifically, an oxygen-species-responsive 3D-printed scaffold with shell-core nanoparticles is designed, which are loaded with an antimicrobial peptide plasmid (LL37 plasmid) and have LL37 grafted on their surface (LL37@ZIF8-LL37). The surface-grafted LL37 directly kills S. aureus and, following entry into cells, the nanoparticles kill intracellular bacteria. Moreover, in vitro and in vivo, following translation of the LL37 plasmid, cells function as factories of the antimicrobial peptide, thereby generating a continuous, prolonged antibacterial effect at the site of infection. This system significantly reduces the abnormal increase in MDSCs within the infected microenvironment, thus relieving the immunosuppressive state and restoring a protective antimicrobial immune response. Hence, this proposed antimicrobial system provides an antimicrobial immune response and a novel strategy for S. aureus-related infections by offering a combined active antimicrobial and immunotherapeutic strategy, thereby significantly reducing the recurrence rate following recovery from implant-associated infections.

Caveolin-1 promotes cancer progression via inhibiting ferroptosis in head and neck squamous cell carcinoma.

BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) is an aggressive disease worldwide. Much progress has been made in exploring mechanisms and improving the therapy of HNSCC, but only a few studies have focused on the role of ferroptosis on HNSCC progression. The current study aimed to reveal the underlining mechanisms that caveolin-1 (CAV1)-ROS (reactive oxygen species)-ferroptosis axis affect the process of HNSCC and discover novo therapeutic targets or strategies. METHODS: The role of CAV1 in ferroptosis was analyzed by FerrDb, and its clinical significance was examined by TCGA dataset of HNSCC. The expressions of caveolin-1 (CAV1) in HNSCC tissues were measured by immunohistochemistry, western blot, and real-time PCR assay. Three siRNA sequences were designed to silence CAV1 mRNA in HNSCC cells. Cell proliferation, colony formation, wound-healing, and transwell assays were used to examine the proliferation, migration, and invasion of cancer cells. ROS evaluation and intracellular Fe2+ content assays were performed to examine the levels of ferroptosis. RESULTS: Through the analysis with published data, CAV1 was found to overexpress in HNSCC than normal tissues, and was one of the vital suppressors of ferroptosis pathway. Our study showed that CAV1 was over expressed in HNSCC tissues and the high level of CAV1 predicted poorer prognosis. Further experiments indicated that CAV1 could inhibit the ferroptosis of cancer cells and promote the proliferation, migration and invasion. CONCLUSIONS: Overexpression of CAV1 in HNSCC inhibited the process of ferroptosis, leading to aggressive phenotypes, as well as worse prognosis. The regulatory pathway of CAV1 and ferroptosis are potential targets for designing diagnostic and combined therapeutic strategies for HNSCC patients.

A quick method of determining the age of 8 years old: Based on the first premolars on Eastern Chinese population.

BACKGROUND: Whether an individual is over 8 years old is of great significance in clinical medicine and law. The Demirjian, Willems and Cameriere methods are one of the most widely used methods of dental age estimation, but neither method was suitable in Eastern China. And all of them are time-consuming for clinical work. A new and time-saving method for judging whether an individual is 8 years old is yet to be proposed on Eastern Chinese population. AIM: The paper proposes a method for judging whether an individual is 8 years old based on the relationship between age and the first premolar maturity index (I1PM), which is related to the measurement of the open apices of the first premolar. SUBJECTS & METHODS: It is a retrospective study where orthopantomograms of 748 children (356 female and 392male) aged 5-13 years were collected and analyzed. Linear regression was used to infer the value of I1PM for eastern China. Furthermore, this method was compared to those based on Demirjian's stages F. RESULTS AND CONCLUSIONS: A cutoff value of I1PM = 0.35 was taken. The sensitivity of this test was 90.0% and specificity was 93.3%. Furthermore, the proportion of individuals with a correct classification was 90.9%. The results of the test showed that Demirjian's stage F performance was lightly better than I1pm < 0.35. However, the sensitivity, specificity, and overall accuracy of both are greater than or equal to 90.0%, and the differences are not very significant.

An improved lentiviral system for efficient expression and purification of ß-defensins in mammalian cells.

ß-Defensins are a family of conserved small cationic antimicrobial peptides with different significant biological functions. The majority of mammalian ß-defensins are expressed in epididymis, and many of them are predicted to have post-translational modifications. However, only a few of its members have been well studied due to the limitations of expressing and purifying bioactive proteins with correct post-translational modifications efficiently. Here we developed a novel Fc tagged lentiviral system and Fc tagged prokaryotic expression systems provided new options for ß-defensins expression and purification. The novel lentiviral system contains a secretive signal peptide, an N-terminal IgG Fc tag, a green fluorescent protein (GFP), and a puromycin selection marker to facilitate efficient expression and fast purification of ß-defensins by protein A magnetic or agarose beads. It also enables stable and large-scale expression of ß-defensins with regular biological activities and post-translational modification. Purified ß-defensins such as Bin1b and a novel human ß-defensin hBD129 showed antimicrobial activity, immuno-regulatory activity, and expected post-translational phosphorylation, which were not found in Escherichia coli (E. coli) in expressed form. Furthermore, we successfully applied the novel system to identify mBin1b interacting proteins, explaining Bin1b in a better way. These results suggest that the novel lentiviral system is a powerful approach to produce correct post-translational processed ß-defensins with bioactivities and is useful to identify their interacting proteins. This study has laid the foundation for future studies to characterize function and mechanism of novel ß-defensins.

Multifunctional nanoplatforms as cascade-responsive drug-delivery carriers for effective synergistic chemo-photodynamic cancer treatment.

Synergistic chemo-photodynamic therapy has garnered attention in the field of cancer treatment. Here, a pH cascade-responsive micellar nanoplatform with nucleus-targeted ability, for effective synergistic chemo-photodynamic cancer treatment, was fabricated. In this micellar nanoplatform, 5-(4-carboxyphenyl)-10,15,20-triphenylporphyrin (Por), a photodynamic therapy (PDT) agent was utilized for carrying the novel anticancer drug GNA002 to construct a hydrophobic core, and cyclic RGD peptide (cRGD)-modified polyethylene glycol (PEG) (cRGD-PEG) connected the cell-penetrating peptide hexaarginine (R6) through a pH-responsive hydrazone bond (cRGD-PEG-N = CH-R6) to serve as a hydrophilic shell for increasing blood circulation time. After passively accumulating in tumor sites, the self-assembled GNA002-loaded nanoparticles were actively internalized into cancer cells via the cRGD ligands. Once phagocytosed by lysosomes, the acidity-triggered detachment of the cRGD-PEG shell led to the formation of R6-coated secondary nanoparticles and subsequent R6-mediated nucleus-targeted drug delivery. Combined with GNA002-induced nucleus-specific chemotherapy, reactive oxygen species produced by Por under 532-nm laser irradiation achieved a potent synergistic chemo-photodynamic cancer treatment. Moreover, our in vitro and in vivo anticancer investigations revealed high cancer-suppression efficacy of this ideal multifunctional nanoplatform, indicating that it could be a promising candidate for synergistic anticancer therapy.

Nuclear-targeted nanocarriers based on pH-sensitive amphiphiles for enhanced GNA002 delivery and chemotherapy.

GNA002, a novel EZH2 inhibitor, exhibits significant anticancer efficiency in solid malignant tumor therapy; however, its poor water solubility and low enrichment at tumor sites limit its clinical application and translation. In this study, an original pH-sensitive nanocarrier (cyclo (RGDyCSH) (cRGD)-poly (ethylene glycol) (PEG)-hydrazine (Hyd)-hexa-arginine (R6)-stearic acid (SA)) was designed to precisely deliver GNA002 into the nuclei of cancer cells. The PEG-modified hydrophilic shell of the spherical GNA002-loaded nanoparticles with a mean size of 143.13 ± 0.20 nm effectively facilitated the passive target of tumor tissues and prolonged the blood circulation time. Meanwhile, cRGD was used as the active targeting ligand, which promoted the accumulation of the nanoparticles in cancer cells via ανß3-receptor-mediated endocytosis. Furthermore, the acidic environment of lysosomes triggered the rupture of the pH-sensitive hydrazine bond and the rapid formation of penetrating peptide R6-shelled secondary nanoparticles, thus enabling the lysosomal escape of the nanoparticles and the ultimate R6-mediated nuclear-targeted delivery of GNA002. Consequently, the nuclear-enriched GNA002 effectively enhanced the cytotoxicity against cancer cells both in vitro and in vivo, thus providing an original and promising drug delivery system for the targeted delivery of GNA002.

A modified dental age assessment method for 5- to 16-year-old eastern Chinese children.

OBJECTIVES: Age estimation is widely applied in the field of orthodontics, pediatric dentistry, and forensic science. Dental age estimation by the radiological method is frequently used because of its convenience and noninvasiveness. However, there are not enough suitable methods for eastern Chinese children. This study aimed to establish a modified formula for eastern Chinese children according to the Demirjian method and then compared the accuracy of the modified method with the Demirjian method and Willems method. MATERIALS AND METHODS: A total of 2367 dental panoramic radiographs from individuals aged 5-16 years of eastern China were collected as samples. Age estimation was conducted using the Demirjian and Willems methods. The polynomial curve fitting method was used to modify the Demirjian method to improve its application to the eastern Chinese children. The paired t test and accuracy ratio were used to compare the applicability of the modified methods with two commonly used methods. RESULTS: The mean chronological age (CA) of the subjects was 11.20 ± 3.29 years for boys and 10.99 ± 3.12 years for girls. The mean difference values between the CA and dental age (DA) (CA-DA) using the Demirjian and Willems methods were 0.73 and 0.7 for boys, respectively, and both 0.79 for girls. The modified method using the polynomial curve fitting presented a smaller underestimation compared with CA for both boys (0.04 years) and girls (0.09 years), which showed a high suitability to Chinese children to some extent. CONCLUSIONS: The Willems method was more accurate in estimating DA compared with the Demirjian method. However, the modified method was more accurate than the two methods; therefore, it can be used in eastern Chinese children. CLINICAL RELEVANCE: It was thought to be a non-invasive, convenient, and efficient method to connect DA and CA. By estimating dental age, pediatrist, and orthodontists can better understand the development of permanent teeth and provide a more accurate orthodontic treatment time and treatment plan to children patients.

Testing the nonlinear equations for dental age evaluation in a population of eastern China.

OBJECTIVE: The study evaluated the validity of the nonlinear equations (Qingdao model) for dental age assessment in an eastern Chinese population. MATERIALS AND METHODS: We studied 1073 digital panoramic radiographs of children aged 11-16 years from a Chinese Han population. Dental ages (DAs) were calculated using the Demirjian and the new model methods. Statistical significance was set at p < 0.05. For each method, differences between the chronological age (CA) and dental age were analyzed by paired t-tests and mean absolute error (MAE). RESULTS: The discrepancies between CA and DA determined by Qingdao model were 0.18 and 0.30 years for males and females, respectively. While using Demirjian method, these differences were and 0.46 and 0.30. The Qingdao model's MAEs between DA and CA were 1.23 and 0.90 years in males and females, respectively. As for the Demirjian method, MAEs were 1.43 and 0.86 years in males and females. CONCLUSIONS: This study showed that the new nonlinear equations were more accurate than the traditional Demirjian method. Especially, the new nonlinear Qingdao model is more competitive in 11-14-year male groups and 15-16-year female groups. We recommend a combined Qingdao model and Demirjian method may reasonably reflect the CAs among children in the eastern Chinese population.

Applicability of newly derived second and third molar maturity indices for indicating the legal age of 16 years in the Southern Chinese population.

In China, forensic age estimation in the living has become increasingly important in these few years. Due to increasing juvenile crimes, the age of 16 years has become the legal age threshold for determining whether the juvenile criminal law or adult criminal law applies to the citizen. This study aimed to assess new cut-off values of the third molar maturity index (I3M) and second molar maturity index (I2M) at the legal age threshold of 16 years in Southern Chinese population and then to compare the applicability of the cut-off values of I3M and I2M we set for discriminating whether a subject is ≥ 16 years. Orthopantomograms (OPGs) of 671 healthy Southern Chinese subjects (332 males and 339 females), aged between 10 and 20 years, were studied. Logistic regression analysis was performed with chronological age (below 16 years and over 16 years) as a dichotomous dependent variable and, I3M, I2M, and sex as predictive variables. The high p-value for sex (p = 0.861) reveals that this factor was not statistically significant in assessing the legal age of 16. The cut-off values of I3M < 0.38 and I2M < 0.03 were identified and used to distinguish between individuals who were or were not aged ≥ 16 years. Compared with a single predictor (I3M) alone, combining I3M and I2M can be more reliable for determining whether an individual is aged more than 16 years. We found that accuracy was 88.52% (95% confidence interval CI, 86.11-90.94%) and sensitivity and specificity were 77.96% (95% CI, 74.82-81.10%) and 97.28% (95% CI, 96.04-98.51%), respectively. The estimated Bayes post-test probability was 99.12% (95% CI, 98.41-99.83%). In conclusion, combining I3M and I2M may be useful in legal and forensic practices to determine the legal age of 16 years in Southern Chinese individuals.

Applicability of the third molar maturity index for assessment of age of majority in Eastern China.

From a legal and penalty point of view, it is essential to conclude if an individual has reached the legal age (also known as age of majority). Although Cameriere's third molar maturity index (I3M) has been used to discriminate between adults and minors, no study has tested the applicability of I3M in the Eastern Chinese population. The aim of this study was to assess the validity of the region-specific cut-off value of I3M for discriminating minors from adults in an Eastern Chinese population. Five hundred fifty-six subjects (276 males and 280 females) aged 14-24 years participated in this study. A logistic regression analysis was conducted by considering the adult and minor ages as dichotomous dependent variables and I3M and sex as predictor variables. The results showed that sex was not statistically significant in distinguishing adults and minors. The receiver operating curve analysis showed the best performance of the cut-off value of I3M < 0.08 in discriminating adults from minors. Furthermore, the diagnostic test showed the proportion of accuracy was 90.22% in males and 86.43% in females. The sensitivity and specificity for males were 88% and 94.06%, respectively, while those for females were 83.71% and 91.18%, respectively. The estimated Bayes post-test probability was 97.18% and 96.01% in males and females respectively. Therefore, I3M < 0.08 may be a useful tool for indicating the legal age in Eastern Chinese population.

Applicability and accuracy of Demirjian and Willems methods in a population of Eastern Chinese subadults.

BACKGROUND: To assess the validity of the Demirjian and Willems dental age estimation methods in a population of Eastern Chinese 11-18-year-old subadults. SAMPLES AND METHODS: A total of 1622 orthopantomograms (787 boys and 835 girls) aged from 11.00 to 18.99 years old from a Chinese Han population were evaluated in the study. Dental ages were calculated using both Demirjian and Willems method. Statistical significance was set at p<0.05 and for each method, discrepancies between chronological age (CA) and dental age (DA) were statistically analyzed by paired t-tests. Special age interval analysis was conducted by Chi-square tests to compare the accuracy of the Demirjian and the Willems methods. RESULTS: Demirjian method underestimated CA 0.66±1.45 y (p<0.001) in boys and 0.62±1.61 y (p<0.001) in girls while Willems method underestimated 0.90±1.32 y (p<0.001) and 0.97±1.56 y (p<0.001) in boys and girls, respectively. The mean absolute error (MAE) value between the CA and the DA (CA-DA) using the Demirjian method and the Willems method were 1.31 and 1.29 y for boys, respectively, and 1.35 and 1.43 y for girls, respectively. CONCLUSIONS: It is suggested that the Demirjian method was found to be more accurate compared with the Willems method. However, the Demirjian method may not be satisfactory for 11-18-year-old children of Eastern China. It is recommended that a modified Demirjian method for better dental age estimation for Eastern Chinese juvenile individuals.