Development and validation of biomarkers related to PANoptosis in osteoarthritis.

OBJECTIVE: Osteoarthritis (OA) is a high-incidence disease of the orthopedic system. However, studies on the molecular mechanisms of OA and pyroptosis, apoptosis, and necroptosis (PANoptosis) at the transcriptome level remain scarce. Therefore, this study purposed to detect biomarkers in OA and explore their relationship to the immune microenvironment. MATERIALS AND METHODS: OA-related expression data was sourced from the Gene Expression Omnibus (GEO) database. Subsequently, differentially expressed analysis and a Venn diagram were performed to obtain differentially expressed PANoptosis-related genes (DEPGs). Furthermore, the least absolute shrinkage and selection operator (LASSO), Support Vector Machine-Recursive Feature Elimination (SVM-RFE), and random forest (RF) were implemented to screen diagnostic genes. Receiver operating characteristic (ROC) curves were performed to verify the diagnostic ability of the diagnostic genes. Next, immune infiltration analysis was performed to find the relationships between differential immune cells (OA vs. normal) and diagnostic genes. Finally, drug prediction analysis was also carried out, and the expression of diagnostic genes was verified in external datasets. RESULTS: A total of 62 DEPGs were identified, which were enriched for regulating apoptotic signaling pathways, tumor necrosis factor (TNF) signaling pathways, and other related pathways. Three feature genes, nuclear factor-kappa-B inhibitor-alpha (NFKBIA), RING finger protein 34 (RNF34), and serine incorporator 3 (SERINC3) were obtained by intersecting genes obtained by the LASSO regression algorithm, SVM algorithm, and RF algorithm and showed excellent diagnostic efficacy with the Area under the curve (AUC) values of individual genes were all greater than 0.7, indicating that the model was more effective. Immuno-infiltration analysis showed that RNF34 was positively correlated with CD56dim natural killer cells, type 17 helper T cells, and NFKBIA was positively correlated with plasmacytoid dendritic cells. Additionally, 12 drugs were predicted by NFKBIA, such as gambogic acid and dioscin. In addition, NFKBIA and SERINC3 were significantly downregulated, and RNF34 was upregulated in OA samples. CONCLUSIONS: Three genes (NFKBIA, RNF34, and SERINC3) related to PANoptosis, were obtained by bioinformatics analysis, which would provide a new direction for the diagnosis and treatment of OA.

[A retrospective analysis of 147 cases of orbital exenteration].

Objective: To investigate the main indications and prognosis of patients undergoing orbital exenteration (OE). Methods: It was a retrospective case series study. The medical records were collected of patients who underwent OE surgery at Beijing Tongren Hospital from January 2007 to December 2021, and their general information, tumor characteristics, lymph node metastasis, diagnosis and treatment, specific surgical techniques, pathological diagnosis and prognosis were analyzed. Kaplan-Meier survival curves were used to evaluate overall survival and recurrence-free survival. Results: A total of 147 patients were included, with a median age of 58 (48, 68) years. Among them, 82 patients (55.8%) were male and 65 (44.2%) were female, and all underwent unilateral OE. Five patients (3.4%) had benign lesions. Among the 142 patients with malignant tumors, conjunctival malignancies (37.3%) were the most common, followed by eyelid (29.6%), orbital (19.0%), and ocular (14.1%) malignancies. Among the 53 patients with conjunctival tumors, 38 (71.7%) had conjunctival melanoma and 14 (26.4%) had squamous cell carcinoma. Among the 42 patients with eyelid malignancies, 19 (45.2%) had sebaceous gland carcinoma and 16 (38.1%) had basal cell carcinoma. Among the 20 patients with ocular malignancies, 10 (50.0%) had choroidal melanoma and 9 (45.0%) had retinoblastoma. The most common pathological types among the 142 patients with malignant tumors were melanoma (51 cases, 35.9%), squamous cell carcinoma (20 cases, 14.1%), sebaceous gland carcinoma (19 cases, 13.4%), and basal cell carcinoma (16 cases, 11.3%). Of the 135 patients included in the survival analysis, all 5 patients with benign lesions were alive at the last follow-up. The median follow-up time for the 130 patients with malignant tumors was 6.9 (2.5, 6.9) years, ranging from 0.2 to 14.0 years. The overall survival rates at 1, 3, and 5 years after surgery were 90.9% (95%CI: 85.8%-96.0%), 68.4% (95%CI: 59.6%-77.2%), and 60.1% (95%CI: 50.5%-69.7%), respectively. The recurrence-free survival rates of patients with and without lymph node metastasis before surgery were 57.6% and 56.7%, respectively (OR=1.062, 95% CI: 0.525-2.148, P=0.864), and the overall survival rates were 61.5% and 57.7%, respectively (OR=1.019, 95% CI: 0.512-2.033, P=0.957), with no significant differences both. The recurrence-free survival rates of patients with melanoma (47 cases) and non-melanoma (83 cases) were 40.4% and 67.5%, respectively (OR=2.576, 95% CI: 1.390-4.775, P<0.001), and the overall survival rates were 42.6% and 67.5%, respectively (OR=2.845, 95% CI: 1.549-5.225, P<0.001), with significant differences both. Conclusions: The primary indications for OE are malignant tumors of the conjunctiva and eyelids, with melanoma being the most common malignant tumor of the conjunctiva. Melanoma patients who undergo OE have a lower survival rate compared to other types of malignant tumors. Nevertheless, even in the presence of tumor lymph node metastasis, patients can achieve a relatively good prognosis through OE.

Raspberry ketone glucoside suppresses melanin synthesis through IL6/JAK1/STAT3 signal pathway.

OBJECTIVE: This study aimed to evaluate the anti-melanogenic activity of raspberry ketone glucoside (RKG) and further explore the specific molecular mechanisms by which RKG affects melanogenesis. MATERIALS AND METHODS: The B16F10 cells model, the mushroom tyrosinase model and the zebrafish model were used to assess the whitening activity of RKG. We subsequently identified possible pathways related to RKG inhibition of melanogenesis by RNA-seq analysis and qRT-PCR on the zebrafish model, and further explored the effects of key genes on the pathway on the melanogenic effect of RKG by using related pathway inhibitors and Tg [mpeg: EGFP] transgenic zebrafish line. RESULTS: RKG could noticeably inhibit melanogenesis in B16F10 cells in vitro and on zebrafish in vivo. The RNA-Seq analysis and the qRT-PCR in zebrafish embryos indicated that the inhibition of melanogenesis by RKG could be achieved by activating JAK1/STAT3 signal pathway and inhibiting the expression levels of the MITFa, TYR, TYRP1a genes directly associated with melanogenesis. The inhibitor tests revealed that the inhibitory effect of the RKG on melanogenesis was restored by the IL6, JAK1/2, and STAT3 inhibitors, specifically STAT3 inhibitor. We further examine the relationship between the JAK1/STAT3 signal pathway and the MITFa. The achieved results indicate that the RKG could activate the zebrafish macrophages via the JAK1, but the inhibition of macrophage activation by loganin did not affect the anti-pigmentation effect of the RKG. CONCLUSIONS: RKG showed remarkable whitening activity on both B16F10 cells in vitro and zebrafish model in vivo. Furthermore, RKG could inhibit melanogenesis by activating the IL6/JAK1/STAT3 pathway, inhibiting the transcriptional activity of MITFa, and its downstream expression levels of the TYR and TYRP1a genes.

[A case of Pfeiffer syndrome caused by FGFR2 gene variation].

A 29-month-old male child with FGFR2 heterozygous missense mutation at birth was diagnosed as Pfeiffer syndrome. He was treating for binocular exophthalmos and exposed keratitis in Beijing Tongren Hospital Affiliated to Capital Medical University. The child had skull fusion (clover head), obvious exophthalmos, deformity of fingers and toes, ankylosis of elbow joint or bony fusion, accompanied by neurological complications and growth retardation; FGFR2 (NM_001144916) gene c.679T>G (thymine>guanine) and p.c227gG(cysteine>glycine) heterozygous missense mutations were found in the the child, and his parents did not carry the same mutation. Pfeiffer syndrome type Ⅱ was diagnosed. Permanent adhesion of eyelid margin was performed under general anesthesia, and the postoperative condition was stable.

MiR-379-5p improved locomotor function recovery after spinal cord injury in rats by reducing endothelin 1 and inhibiting astrocytes expression.

OBJECTIVE: The aim of this study was to investigate the effect of microRNA-214-5p (miR-214-5p) on spinal cord injury (SCI) and to explore the mechanism of action in pathophysiological relevance. MATERIALS AND METHODS: The model of SCI was successfully established in rats aged 6-8 weeks. The levels of the locomotor function recovery in rats of the miR-379-5p group and SCI group were detected one month later by Basso-Beattie-Bresnahan (BBB) locomotor rating scale. Biochemical indexes were measured by Western blotting and real time-PCR, respectively. In addition, rat astrocytes were cultured to verify the effect of miR-379-5p on activated astrocytes in vitro. RESULTS: Compared with the SCI group, the rats in the miR-379-5p group showed prominent improvement in the locomotor function in vivo. MiR-379-5p attenuated the activation of astrocytes and significantly suppressed the expressions of the nerve growth inhibitors. Furthermore, the down-regulation of endothelin-1 (ET-1) ameliorated the spinal cord ischemia, thereby reducing apoptosis and oxidative stress. Compared with the pentylenetetrazol (PTZ) group, ET-1 and chondroitin sulfate poly-glycoprotein (CSPG) in miR-379-5p group decreased significantly in the astrocytes transfected with miR-214-5p in vitro. CONCLUSIONS: MiR-379-5p retarded the neurofilament regeneration block effect by inhibiting endothelin 1 and the expression of the astrocytes after SCI. Furthermore, it might relieve nerve structure destruction, resist oxidative stress, and inhibit apoptosis, eventually promoting functional recovery.

Effects of anodic titanium dioxide nanotubes of different diameters on macrophage secretion and expression of cytokines and chemokines.

OBJECTIVES: To investigate effects of TiO2 nanotubes of different diameters on J744A.1 macrophage behaviour, secretion and expression of pro-inflammatory cytokines and chemokines. MATERIALS AND METHODS: Macrophage-like J744A.1 cells were cultured on three types of Ti surface: mechanically polished titanium plus 30 and 80 nm TiO2 nanotube surfaces, for 4, 24 and 48 h. Macrophage adhesion and proliferation were assessed using CCK-8 assay. Levels of pro-inflammatory cytokines (TNF-α, IL-1ß and IL-6) and chemokines (MCP-1 and MIP-1α) secreted into the supernatant were measured using the Cytometric Bead Arrays kit. TNF-α, MCP-1 and MIP-1α gene expression were quantitatively analysed by real-time PCR. RESULTS: These show that TiO2 nanotube surfaces, especially of 80 nm TiO2 nanotube, benefited macrophage adhesion and proliferation, and reduced protein secretion and mRNA expression of TNF-α, MCP-1 and MIP-1α. IL-1ß and IL-6 were undetectable on all the surfaces at all times. CONCLUSIONS: TiO2 nanotube surfaces, especially of 80 nm TiO2 nanotube, reduced inflammatory response in vitro, which might be part of a basis for rapid osseointegration in implants with TiO2 nanotube surfaces in animal studies.

Association between rs11001553 of DKK1 and non-syndromic tooth agenesis in the Chinese Han population.

Tooth agenesis is one of the most common anomalies of human dentition. Recent genetic studies have provided information regarding a number of genes related to both syndromic and non-syndromic forms of hypodontia. In a previous study, we found that polymorphism in rs11001553 of DKK1 was associated with hypodontia in the Chinese Han population. In this study, we extended this investigation to 89 individuals diagnosed with sporadic non-syndromic oligodontia (40 males and 49 females). These individuals were analyzed with 268 subjects (123 males and 145 females) diagnosed with non-syndromic hypodontia and 190 healthy control subjects (99 males and 91 females). DNA was obtained from whole blood or saliva samples and genotyping was performed by matrix-assisted laser desorption ionization-time of flight mass spectrometry. Significant differences were observed in the allele and genotype frequencies of rs11001553 of DKK1. These data demonstrated an association between rs11001553 of DKK1, a tooth development-associated gene, and non-syndromic tooth agenesis in Chinese Han individuals. This information may provide further understanding of the molecular mechanisms of tooth agenesis. Furthermore, DKK1 can be regarded as a marker gene for the risk of tooth agenesis.

Family-based association analysis of S100A8 genetic polymorphisms with aggressive periodontitis.

BACKGROUND AND OBJECTIVE: It is known that S100A8, a member of the S100 calcium-binding protein family, is associated with inflammatory diseases, including periodontitis. Our previous population-based study found an association between two polymorphisms, rs3795391 (A > G) and rs3806232 (A > G), in the upstream region of the S100A8 gene and aggressive periodontitis (AgP) in Chinese people. Based on those results, this investigation set out to analyze and corroborate whether the association also exists within families. MATERIAL AND METHODS: Two hundred and four subjects from 73 nuclear families were recruited. All probands and their relatives were diagnosed according to the 1999 classification of periodontal diseases. Anticoagulated peripheral blood samples were collected from all the subjects, and DNA was extracted. The two single nucleotide polymorphisms (SNPs; rs3795391 and rs3806232) were detected and analyzed by standard polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay. Analysis of genotype/allele was performed by Family-Based Association Test (FBAT) software ( http://www.biostat.harvard.edu/~fbat/default.html). RESULTS: There was a statistically significant association of the SNP rs3795391 with AgP in the additive genetic model (chi(2) = 3.9836, d.f. = 1, p = 0.0459). Allele A showed significantly preferential transmission to the AgP affected individuals (Z = 1.996, p = 0.0459). The other SNP, rs3806232, showed no significant results in all models. CONCLUSIONS: This family-based association study supports the previous findings that SNP rs3795391 (A > G) of the S100A8 gene might contribute to AgP susceptibility. This is, to our knowledge, the first investigation about AgP using FBAT in genetic analysis.

LEDGFp52 controls rat retinal ganglion cell neurite growth in culture and regulates specific neuronal growth-associated genes and protein production.

We investigated the regulation of primary neurite growth and expression of specific growth-associated genes by lens epithelium-derived growth factor (LEDGF) in rat retinal ganglion cells (RGCs). A pAd-LEDGFp52 adenovirus vector and a siRNA-LEDGFp52 eucaryotic expression vector were transfected into cultured RGCs. Transfection with pAd-LEDGFp52 significantly increased the number of neurites and their lengths compared with untransfected control RGCs. The expression of growth associated protein 43 (GAP43), microtubule-associated protein 2 (MAP2), and low-molecular-weight neurofilament (NF-L) genes and proteins were also significantly up-regulated. In contrast, the introduction of siRNA-LEDGFp52 significantly decreased the number and length of neurites, and significantly down-regulated the expression GAP43, NF-L and MAP2 genes and proteins compared with controls. Our findings suggest that LEDGFp52 might act as a dendritic arborization gene as well as an axonal elongation gene in RGCs and that it might be beneficial to the functional recovery of regenerating RGCs.

PTCH germline mutations in Chinese nevoid basal cell carcinoma syndrome patients.

OBJECTIVES: PTCH, the human homologue of the Drosophila segment polarity gene, patched, has been identified as the gene responsible for nevoid basal cell carcinoma syndrome. The aim of this study was to investigate PTCH gene mutation in Chinese patients with nevoid basal cell carcinoma syndrome. MATERIALS AND METHODS: DNA was isolated from both odontogenic keratocyst tissue and peripheral blood of five patients with syndrome and one patient with only multiple odontogenic keratocysts, and mutational analysis of the PTCH gene performed by direct sequencing after amplification of all 23 exons by polymerase chain reaction (PCR). RESULTS: A previously reported germline mutation (c.2619C>A) was identified in two familial cases involving the mother and the daughter, with the mother also carrying a novel somatic mutation (c.361_362insGAGC). Three novel germline PTCH mutations (c.1338_1339insGCG, c.331delG and c.1939A>T) were detected in three unrelated patients with syndrome. The patient with multiple odontogenic keratocysts who failed to fulfill the diagnostic criteria of the syndrome also carried a novel germline mutation (c.317T>G). CONCLUSION: The frequent germline PTCH mutations detected in our series provide further evidence for the crucial role of PTCH in the pathogenesis of nevoid basal cell carcinoma syndrome in Chinese.

The application of genomic and proteomic technologies in predictive, preventive and personalized medicine.

The long asymptomatic period before the onset of chronic diseases offers good opportunities for disease prevention. Indeed, many chronic diseases may be preventable by avoiding those factors that trigger the disease process (primary prevention) or by use of therapy that modulates the disease process before the onset of clinical symptoms (secondary prevention). Accurate prediction is vital for disease prevention so that therapy can be given to those individuals who are most likely to develop the disease. The utility of predictive markers is dependent on three parameters, which must be carefully assessed: sensitivity, specificity and positive predictive value. Specificity is important if a biomarker is to be used to identify individuals either for counseling or for preventive therapy. However, a reciprocal relationship exists between sensitivity and specificity. Thus, successful biomarkers will be highly specific without sacrificing sensitivity. Unfortunately, biomarkers with ideal specificity and sensitivity are difficult to find for many diseases. One potential solution is to use the combinatorial power of a large number of biomarkers, each of which alone may not offer satisfactory specificity and sensitivity. Recent technological advances in genetics, genomics, proteomics, and bioinformatics offer a great opportunity for biomarker discovery. The newly identified biomarkers have the potential to bring increased accuracy in disease diagnosis and classification, as well as therapeutic monitoring. In this review, we will use type 1 diabetes (T1D) as an example, when appropriate, to discuss pertinent issues related to high throughput biomarker discovery.

PTCH mutations in sporadic and Gorlin-syndrome-related odontogenic keratocysts.

Odontogenic keratocysts are relatively common lesions that may occur in isolation or in association with nevoid basal cell carcinoma syndrome (or Gorlin syndrome). The PTCH gene has been reported to be associated with Gorlin syndrome. We investigated 10 cases of non-syndromic keratocysts and two other cases associated with Gorlin syndrome, looking for PTCH mutations. Four novel and 1 known PTCH mutations were identified in five individual patients. Of the 5 mutations identified, 2 were germ-line mutations (2619C>A; 1338_1339insGCG) in 2 cysts associated with Gorlin syndrome, and 3 were somatic mutations (3124_3129dupGTGTGC; 1361_1364delGTCT; 3913G>T) in 3 non-syndromic cysts. This report describes PTCH mutations in both non-syndromic and Gorlin-syndrome-related odontogenic keratocysts in Chinese patients, and suggests that defects of PTCH are associated with the pathogenesis of syndromic as well as a subset of non-syndromic keratocysts.

Antiapoptotic and antioxidant effects of rosmarinic acid in astrocytes.

The protective effects of rosmarinic acid (RA), a polyphenolic compound, on apoptosis induced by hydrogen peroxide in astrocytes were studied. Pretreating cells with RA significantly increased cell viability and decreased apoptosis rate induced by H2O2. The antiapoptotic effect of RA was further confirmed by increase of mitochondrial membrane potential and inhibition of caspase-3 activity. RA also attenuated cellular oxidative stress by decreasing the amount of reactive oxygen species and malondialdehyde. Results clearly show that RA was able to attenuate H2O2-induced cell injury by its antiapoptotic and antioxidant activity.

[Experiments with the technology for manufacturing oral liquid of wuji jing].

An improved technological process for manufacturing the oral liquid of Wuji Jing (the essence of flesh of the black-boned chicken) is presented. In the present technique we adopted a hydrolytic procedure instead of extraction by boiling, as used in the original technology. We use some proteinase as hydrolytic agent.