A novel LINC02321 promotes cell proliferation and decreases cisplatin sensitivity in bladder cancer by regulating RUVBL2.

Bladder cancer (BC) has a high incidence and is prone to recurrence. In most instances, the low 5-year survival rate of advanced BC patients results from postoperative recurrence and drug resistance. Long noncoding RNAs (lncRNAs) can participate in numerous biological functions by regulating the expression of genes to affect tumorigenesis. Our previous work had demonstrated that a novel lncRNA, LINC02321, was associated with BC prognosis. In this study, A high expression of LINC02321 was found in BC tissues, which was associated with poor prognosis in patients. LINC02321 promoted both proliferation and G1-G0 progression in BC cells, while also inhibited sensitivity to cisplatin. Mechanistically, LINC02321 can bind to RUVBL2 and regulate the expression levels of RUVBL2 protein by affecting its half-life. RUVBL2 is involved in the DNA damage response. The potential of DNA damage repair pathways to exert chemosensitization has been demonstrated in vivo. The rescue experiment demonstrated that RUVBL2 overexpression can markedly abolish the decreased cell proliferation and the increased sensitivity of BC cells to cisplatin caused by LINC02321 knockdown. The results indicate that LINC02321 functions as an oncogene in BC, and may serve as a novel potential target for controlling BC progression and addressing cisplatin resistance in BC therapy.

Evaluating the efficacy of recombinant human growth factors in scar remodelling for patients with facial soft tissue injuries.

Facial soft tissue injuries, often resulting in scarring, pose a challenge in reconstructive and aesthetic surgery due to the need for functional and aesthetic restoration. This study evaluates the efficacy of recombinant human growth factors (rhGFs) in scar remodelling for such injuries. A retrospective evaluation was conducted from January 2020 to January 2023, involving 100 patients with facial soft tissue injuries. Participants were divided equally into a control group, receiving standard cosmetic surgical repair, and an observation group, treated with rhGFs supplemented cosmetic surgery. The study assessed scar characteristics (pigmentation, pliability, vascularity, height), hospital stay duration, tissue healing time, complication rates and patient satisfaction. The observation group demonstrated significant improvements in all scar characteristics, with notably better pigmentation, pliability, vascularity and height compared with the control group. The rhGF treatment also resulted in reduced hospital stay duration and faster tissue healing. Notably, the total complication rate was significantly lower in the observation group (10%) compared with the control group (34%). Additionally, patient satisfaction levels were higher in the observation group, with 98% combined satisfaction compared with 76% in the control group. The application of rhGFs in treating facial soft tissue injuries significantly enhances scar remodelling, expedites healing, reduces complications and improves patient satisfaction. These findings establish rhGFs as a valuable tool in the management of facial soft tissue injuries, highlighting their potential in improving both functional and aesthetic outcomes.

HGFIN deficiency exacerbates spinal cord injury by promoting inflammation and cell apoptosis through regulation of the PI3K/AKT signaling pathway.

BACKGROUND: Spinal cord injury (SCI) is a devastating neurological disease characterized by neuroinflammation and neuronal apoptosis. The PI3K/AKT signaling pathway is related to the pathological process of SCI. Hematopoietic growth factor inducible neurokinin-1 type (HGFIN) is a transmembrane glycoprotein that exerts neuroprotective actions in various neurodegenerative diseases. However, the potential role and mechanism of HGFIN in the development of SCI are still unclear. OBJECTIVES: To investigate the effect of HGFIN on inflammation and neuronal apoptosis as well as the underlying mechanism in SCI. MATERIAL AND METHODS: A rat model of SCI was established, and Basso-Beattie-Bresnahan (BBB) motor function assay was performed to detect motor function. Expression of HGFIN was measured at 7 days after injury by western blot and immunofluorescence. An HGFIN-shRNA-carrying lentivirus was injected into the injury site to block the expression of HGFIN. The effects of HGFIN on neuronal apoptosis and the PI3K/AKT pathway were analyzed by TUNEL staining and immunofluorescence. The Iba-1 expression and the levels of pro-inflammatory cytokines were measured in spinal cord tissues by immunofluorescence staining and real-time polymerase chain reaction (PCR) analysis. RESULTS: The SCI rats showed increased expression of HGFIN in spinal cord tissues. The HGFIN deficiency aggravated SCI lesions, as evidenced by decreased BBB scores. At 7 days post-injury, HGFIN knockdown promoted neuronal apoptosis, accompanied by the increased expression level of the apoptosis effector cleaved caspase-3 and cleaved PARP, and decreased anti-apoptotic protein Bcl-2 expression. Moreover, HGFIN knockdown aggravated the inflammation process, indicated by increased Iba1-positive cells. The HGFIN knockdown increased the production of pro-inflammatory cytokines including IL-1ß, TNF-α and IL-6. Further analysis revealed that HGFIN deficiency reduced the activation of the PI3K/AKT pathway in spinal cord tissue after injury. CONCLUSIONS: Lentivirus-mediated downregulation of HGFIN exacerbates inflammation and neuronal apoptosis in SCI by regulating the PI3K/AKT pathway, and provides clues for developing novel therapeutic approaches and targets against SCI.

Sperm induce macrophage extracellular trap formation via phagocytosis-dependent mechanism.

Infertility is a public health concern worldwide. Asthenozoospermia is a common cause of male infertility and is characterized by decreased motility. Sperm motility ensures that sperm migrate to complete fertilization. Macrophages are an essential component of innate immunity in the female reproductive tract. Macrophage extracellular traps are induced by various microorganisms to capture and mediate the clearance of microorganisms. The relationship between sperm and macrophage extracellular traps is unclear. The human monocyte leukemia (THP-1) cells differentiated by phorbol myristate acetate (PMA) are widely used as surrogate of human macrophages. This study investigated sperm-induced macrophage extracellular trap formation and clarified some of the mechanisms affecting macrophage extracellular trap production. Sperm-induced macrophage extracellular traps were visualized and components of macrophage extracellular traps were identified by immunofluorescence analyses and scanning electron microscopy. By inhibiting macrophage extracellular trap production and macrophage phagocytosis, the relationship between macrophage phagocytosis and macrophage extracellular trap production was analyzed. Sperm could trigger PMA-differentiated THP-1 macrophages to produce extracellular traps. Sperm-triggered macrophage extracellular traps are dependent on phagocytosis and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Sperm from asthenozoospermia donors are more likely to be phagocytosed by macrophages than sperm from healthy donors, which induce more macrophage extracellular trap release. These data confirm the phenomenon and partial mechanism of sperm-induced macrophage extracellular trap formation in vitro. These may partly provide evidence to explain the mechanisms of clearing abnormally morphological or hypomotile sperm in the female reproductive tract and the rationale for the decreased probability of successful fertilization in asthenozoospermia.

Single-dose rAAV5-based vaccine provides long-term protective immunity against SARS-CoV-2 and its variants.

The COVID-19 pandemic, caused by the SARS-CoV-2 virus and its variants, has posed unprecedented challenges worldwide. Existing vaccines have limited effectiveness against SARS-CoV-2 variants. Therefore, novel vaccines to match mutated viral lineages by providing long-term protective immunity are urgently needed. We designed a recombinant adeno-associated virus 5 (rAAV5)-based vaccine (rAAV-COVID-19) by using the SARS-CoV-2 spike protein receptor binding domain (RBD-plus) sequence with both single-stranded (ssAAV5) and self-complementary (scAAV5) delivery vectors and found that it provides excellent protection from SARS-CoV-2 infection. A single-dose vaccination in mice induced a robust immune response; induced neutralizing antibody (NA) titers were maintained at a peak level of over 1:1024 more than a year post-injection and were accompanied by functional T-cell responses. Importantly, both ssAAV- and scAAV-based RBD-plus vaccines produced high levels of serum NAs against the circulating SARS-CoV-2 variants, including Alpha, Beta, Gamma and Delta. A SARS-CoV-2 virus challenge showed that the ssAAV5-RBD-plus vaccine protected both young and old mice from SARS-CoV-2 infection in the upper and lower respiratory tracts. Whole genome sequencing demonstrated that AAV vector DNA sequences were not found in the genomes of vaccinated mice one year after vaccination, demonstrating vaccine safety. These results suggest that the rAAV5-based vaccine is safe and effective against SARS-CoV-2 and several variants as it provides long-term protective immunity. This novel vaccine has a significant potential for development into a human prophylactic vaccination to help end the global pandemic.

Factors Associated with Pulmonary Function Changes in Patients Undergoing Microwave Ablation for Pulmonary Ground-Glass Nodules.

Purpose: Microwave ablation has become an alternative treatment for pulmonary ground-glass nodules (GGN) and is widely accepted by clinicians. However, its effect on lung function remains unknown. Therefore, this retrospective study aimed to explore pulmonary function changes and associated risk factors in patients undergoing computed tomography (CT)-guided microwave ablation (MWA) for treating pulmonary GGN. Materials and Methods: Thirty-five patients diagnosed with pulmonary GGN on thin-layer chest CT and enhanced CT were examined. Patients unable or unwilling to undergo thoracoscopic surgery underwent CT-guided simultaneous percutaneous core needle biopsy and MWA. Pulmonary function tests (PFT) were performed before ablation and 3 days and 6 months post-ablation. Forced expiratory volume in one second (FEV1), FEV1%, forced vital capacity (FVC), maximal voluntary ventilation (MVV), and peak expiratory flow (PEF) values pre- and post-MWA were analysed. Linear regression analysis was used to examine the correlation between ablation volume and changes in PFT findings 3 days post-ablation. Associations between patient characteristics, rates of postoperative complications, and PFT findings were analysed. Results: Forty-eight lesions were completely ablated and examined intraoperatively. There were significant differences in pre- and post-operative PFT findings on day 3 but not at 6 months. The mean ablation volume after 3 days of 11.4 ± 6.3 cm3 was positively correlated with changes in FEV1, MVV, and PEF values. Patients' age (mean, 59.4 ± 13.0 years) positively correlated with changes in PEF values. The rates of change in FVC and MVV values were significantly higher with multiple pulmonary nodules than with isolated pulmonary nodule. PFT findings were similar between patients who experienced or did not experience complications (eg, pneumothorax and pleural effusion). Conclusions: Pulmonary function could be impaired shortly after MWA. PFT findings may correlate with age, ablation volume, and number of ablated lesions. In most patients, pulmonary function returned to the preoperative state after 6 months.

An enzyme-responsive and NIR-triggered lipid-polymer hybrid nanoplatform for synergistic photothermal/chemo cancer therapy.

A combination of photothermal therapy (PTT) and chemotherapy is an emerging therapeutic strategy with promising clinical prospects in cancer treatment. Despite the huge progress achieved in the past years, a number of obstacles still hamper the therapeutic efficacy of this synergistic modality such as uneven heat distribution, lack of targetability of anti-cancer agents and dosage-related side effects. Thus, developing a nanoplatform for targeted drug delivery against cancer is of great necessity. Herein, a lipid-polymer hybrid nanosystem (LP/ID) based on polyethyleneimine (PEI)-lecithin-polyethylene glycol (PEG) was fabricated to co-load indocyanine green (ICG) and dichloroacetate (DCA) for combined photothermal/chemotherapy. DCA and ICG were linked to the PEI backbone to form a dense hydrophobic core through amide bonds and electrostatic interactions, which increased the payload of DCA and ICG as well as achieved enzyme-responsive drug release because of the overexpressed amidase in tumor cells. Lecithin and DSPE-PEG2000 self-assembled around the hydrophobic complexes to obtain prolonged blood circulation and attenuated systemic toxicity of the hybrid nanosystem. The prepared LP/ID exhibited favourable stability in a physiological environment, good tumor imaging properties, and satisfactory photothermal/chemotherapeutic performance. Moreover, LP/ID could also enhance the cellular uptake and tumor retention capacity in comparison with free drug administration. Notably, by co-loading two therapeutic agents with different anti-cancer mechanisms, an obvious inhibitory effect on tumor growth was observed with negligible damage to normal tissues and organs because of the synergistic photothermal/chemotherapy effect, indicating the great potential of LP/ID as a robust nanoplatform for cancer treatment.

A long noncoding RNA GTF2IRD2P1 suppresses cell proliferation in bladder cancer by inhibiting the Wnt/ß­catenin signaling pathway.

Background: There is growing evidence that long non-coding RNAs (LncRNAs) are key in the development of a variety of human tumors. However, the role of lncRNA GTF2IRD2P1 has not been well studied in cancer. The impact of GTF2IRD2P1 on the biological function and clinical relevance in bladder cancer is largely unknown. This study aimed to investigate the biological role of GTF2IRD2P1 in bladder evolution and carcinogenesis. Methods: We used bioinformatics to obtain the lncRNA GTF2IRD2P1 from bladder urothelial carcinoma (BLCA) in The Cancer Genome Atlas (TCGA) database. The expression of lncRNA GTF2IRD2P1 was detected by qRT-PCR. The CCK8 assay and flow cytometry were used to detect the lncRNA GTF2IRD2P1 function on the proliferation of bladder cancer cells. A western blot was used to calculate the protein level of cell cycle proteins and Wnt signaling pathway proteins. The effect of lncRNA GTF2IRD2P1 on tumorigenesis of bladder cancer was confirmed by a xenograft nude mouse model. Results: GTF2IRD2P1 expression was found to be lower in both human bladder cancer tissues and cell lines (UM-UC-3, RT4, and 5637), and elevated in T24 compared to the corresponding normal controls. GTF2IRD2P1 expression was also enhanced after transfection of UM-UC-3 cells with the overexpression vector. Meanwhile, overexpression of GTF2IRD2P1 inhibited the proliferation of UM-UC-3 and prolonged the cell cycle. The silencing of GTF2IRD2P1 significantly increased the proliferation and shortened the cell cycle of T24 cells and induced Wnt signaling activity to promote the progression of bladder cancer. Similarly, the transplanted tumor nude mouse model demonstrated that silencing GTF2IRD2P1 strengthens the progression of bladder cancer by targeting the Wnt signaling pathway.

Alleviation of doxorubicin-induced cardiomyocyte death through miR-147-y-mediated mitophagy.

Doxorubicin (DOX) is a commonly used antitumor drug. However, it may cause severe cardiotoxicity, apoptosis being a major change. A recent report indicates that miR-147 expression is decreased in the myocardium of a myocardial infarction model, suggesting a potential role of this miRNA in DOX-induced cardiomyocyte toxicity. In this study, freshly isolated neonatal pig cardiomyocytes were used; following transfection of a miR-147-y mimic, the cell death induced by DOX was alleviated, represented by augmented mitophagy [indicated by a decrease in P62, and increases in LC3, PINK1, parkin mRNA, LC3Ⅱ/Ⅰ, beclin-1, PINK1, and parkin including p-parkin (Ser65) protein expression], prohibited cell apoptosis as determined by TUNEL staining, and the suppression of caspase-3 transcription and cleaved caspase-3 translation. In cells transfected with an miR-147-y inhibitor, DOX-induced mitophagy was decreased, while apoptosis was increased. Additionally, RAPTOR gene silencing in cardiomyocytes exposed to DOX increased the rate of mitophagy and decreased that of apoptosis as compared with the treatment with DOX alone. Moreover, RAPTOR overexpression downregulated the rate of mitophagy and increased that of apoptosis in cells exposed to DOX. RAPTOR was confirmed as the target gene of miR-147-y based on the results of luciferase reporter gene assays and the opposite effects of the miR-147-y mimic and miR-147-y inhibitor on RAPTOR expression. In summary, our study suggests that miR-147-y mediates DOX-induced cardiomyocyte mitophagy while suppresses apoptosis by targeting RAPTOR, thus playing a protective role in DOX-induced cardiomyocyte damage.

Human CYP enzyme-activated genotoxicity of 2,2',4,4'-tetrabromobiphenyl ether in mammalian cells.

Polybrominated biphenyl ethers (PBDEs) are a group of persistent organic pollutants with endocrine-disrupting, neurotoxic, tumorigenic and DNA-damaging activities. They are hydroxylated by human liver microsomal CYP enzymes, however, their mutagenicity remains unknown. In this study, 2,2',4,4'-tetrabromobiphenyl ether (BDE-47, relatively abundant in human tissues) was investigated for micronuclei induction and DNA damage in mammalian cells. The results indicated that BDE-47 up to 80 µM under a 6 h/18 h (exposure/recovery, covering 2 cell cycles) regime did not induce micronuclei in V79-Mz and V79-derived cell lines expressing human CYP1A1 or 1A2, while it was moderately positive in human CYP2B6-, 2E1-and 3A4-expressing cell lines (V79-hCYP2B6, V79-hCYP2E1-hSULT1A1 and V79-hCYP3A4-hOR, respectively). Following 24 h exposure, BDE-47 induced micronuclei in V79-hCYP2E1-hSULT1A1 and V79-hCYP3A4 cells at increased potencies. In the human hepatoma (HepG2) cells BDE-47 (48 h exposure) was inactive up to 40 µM, however, pretreatment of the cells with ethanol (0.2%, v:v, inducer of CYP2E1) or rifampicin (10 µM, inducer of CYP3A4) led to significant micronuclei formation by BDE-47; pretreatment with bisphenol AF (100 nM) also potentiated BDE-47-induced micronuclei formation (which was blocked by a CYP2E1 inhibitor trans-1,2-dichloroethylene or a CYP3A inhibitor (ketoconazole). Immunofluorescent staining of centromere protein B with the micronuclei formed by BDE-47 in HepG2 cells pretreated with ethanol or rifampicin demonstrated selective formation of centromere-containing micronuclei. The increased phosphorylation of both histones H2AX and H3 in HepG2 by BDE-47 also indicated an aneugenic potential. Therefore, this study suggests that BDE-47 is an aneugen activated by several human CYP enzymes.

Aurantio-obtusin induces hepatotoxicity through activation of NLRP3 inflammasome signaling.

Aurantio-obtusin (AO) is a major anthraquinone (AQ) compound derived from Cassiae semen (CS). Although pharmacological studies have shown that the CS extracts can serve as effective agents in preclinical and clinical practice, AQ-induced hepatotoxicity in humans has attracted widespread attention. To explore whether AO induces hepatotoxicity and its underlying mechanisms, we exposed larval zebrafish and mice to AO. We found that AO delayed yolk sac absorption, and increased liver area and inflammation in the larval zebrafish. This inflammation was manifested as an increase in liver neutrophils and the up-regulated mRNA expression of interleukin-6 (Il-6) and tumor necrosis factor-α (Tnf-α) in the larval zebrafish. Furthermore, a pharmacokinetics study showed that AO was quickly absorbed into the blood and rapidly metabolized in the mice. Of note, AO induced hepatotoxicity in a gender-dependent manner, characterized by liver dysfunction, increased hepatocyte necrosis with inflammatory infiltration, and up-regulated mRNAs of Il-6, Tnf-α and monocyte chemotactic protein 1(Mcp1) in the female mice after 28-day oral administration. It also highlighted that AO triggered NOD-like receptor protein (NLRP) signaling in the female mice, as evidenced by the increased NLRP3, Caspase-1, pro-IL-1ß, IL-1ß and IL-18. Finally, we found that AO led to a significant increase in potassium calcium-activated channel, subfamily N, member 4 (KCNN4) and reactive oxygen species (ROS) levels, along with decreased nuclear factor kappa B p65 (NF-κB p65), in the female mouse livers. In conclusion, AO induced hepatotoxicity by activating NLRP3 inflammasome signaling, at least in part, through increased KCNN4 and ROS production, and NF-κB inhibition.

Black Phosphorus, an Emerging Versatile Nanoplatform for Cancer Immunotherapy.

Black phosphorus (BP) is one of the emerging versatile nanomaterials with outstanding biocompatibility and biodegradability, exhibiting great potential as a promising inorganic nanomaterial in the biomedical field. BP nanomaterials possess excellent ability for valid bio-conjugation and molecular loading in anticancer therapy. Generally, BP nanomaterials can be classified into BP nanosheets (BPNSs) and BP quantum dots (BPQDs), both of which can be synthesized through various preparation routes. In addition, BP nanomaterials can be applied as photothermal agents (PTA) for the photothermal therapy (PTT) due to their high photothermal conversion efficiency and larger extinction coefficients. The generated local hyperpyrexia leads to thermal elimination of tumor. Besides, BP nanomaterials are capable of producing singlet oxygen, which enable its application as a photosensitizer for photodynamic therapy (PDT). Moreover, BP nanomaterials can be oxidized and degraded to nontoxic phosphonates and phosphate under physiological conditions, improving their safety as a nano drug carrier in cancer therapy. Recently, it has been reported that BP-based PTT is capable of activating immune responses and alleviating the immunosuppressive tumor microenvironment by detection of T lymphocytes and various immunocytokines, indicating that BP-based nanocomposites not only serve as effective PTAs to ablate large solid tumors but also function as an immunomodulation agent to eliminate discrete tumorlets. Therefore, BP-mediated immunotherapy would provide more possibilities for synergistic cancer treatment.

Downregulation of PART1 Inhibits Proliferation and Differentiation of Hep3B Cells by Targeting hsa-miR-3529-3p/FOXC2 Axis.

BACKGROUND: Long noncoding RNAs (lncRNAs) are an important subtype of noncoding RNAs (ncRNAs) and microRNA sponges regulate protein-coding gene expression. The lncRNA prostate androgen-regulated transcript 1 (PART1) was implicated in the process of several cancer pathogeneses. However, studies on the regulation of PART1 expression and its mechanism in liver cancer are lacking. METHODS: qRT-PCR and western blot were used to detect PART1 levels in liver cancer serums and cell lines. Cell proliferation, migration, and invasion were detected using CCK8 assays, cell clones, and transwell assays. Interaction between PART1 and miR-3529-3p and forkhead box protein C2 (FOXC2) was confirmed using dual-luciferase reporter assays. RESULTS: We revealed that expression levels of PART1 and FOXC2 are significantly upregulated and the miR-3529-3p expression level significantly decreases in the serum while high expression level of PART1 is positively associated with tumour size, BCLC stage, and TNM stage. shRNA of PART1 can significantly reduce the ability of cell migration and invasion by regulating AKT signalling associated with the reduction of MMP-2 and MMP-9 protein expression. Dual-luciferase reporter assays showed that PART1 can sponge miR-3529-3p, which targets FOXC2 in liver cancer cells. The promoting or suppressing effect of PART1 for Hep3B cell proliferation, invasion, and migration is revised by miR-3529-3p mimics and inhibitors. CONCLUSION: Results showed that downregulation of PART1 can partially inhibit proliferation and differentiation by targeting hsa-miR-3529-3p/FOXC2 axis.

Caveolin-1 Knockdown Decreases SMMC7721 Human Hepatocellular Carcinoma Cell Invasiveness by Inhibiting Vascular Endothelial Growth Factor-Induced Angiogenesis.

Background: Recently, several studies have demonstrated that caveolin-1 overexpression is involved in apoptosis resistance, angiogenesis, and invasiveness in hepatocellular carcinoma (HCC). However, the mechanisms underlying caveolin-1-mediated tumor progression remain unclear. Methodogy. Lentiviral vectors were used to construct caveolin-1 small interfering RNA- (siRNA-) expressing cells. Secreted VEGF levels in SMMC7721 cells were evaluated by enzyme-linked immunosorbent assay (ELISA). SMMC7721 cell proliferation, cycle, apoptosis, and invasiveness were detected by MTT, flow cytometry, Annexin V-FITC/PI, and invasion assay, respectively. Phospho-eNOS levels in human umbilical vein endothelial cells (HUVECs) cocultured with SMMC7721 cell supernatants were analyzed by Western blot. Capillary-like tubule formation assay was performed to analyze endothelial tubular structure formation in HUVECs treated with supernatants from caveolin-1 siRNA-expressing SMMC7721 cells. SMMC7721 implantation and growth in nude mice were observed. Angiogenesis in vivo was analyzed by immunohistochemical angiogenesis assay. Results: Caveolin-1 siRNA-expressing SMMC7721 cells secreted reduced levels of VEGF. Caveolin-1 RNAi also caused an inhibition of SMMC7721 cell proliferation and cell cycle progression that was accompanied by increased apoptosis. Supernatants from caveolin-1 siRNA-expressing SMMC7721 cells inhibited cell cycle progression and decreased phospho-eNOS levels in HUVECs. Endothelial tubular structure formation in HUVECs treated with supernatants from caveolin-1 siRNA-expressing SMMC7721 cells was considerably reduced. Caveolin-1 siRNA-expressing SMMC7721 cells also showed reduced tumorigenicity and angiogenesis induction in vivo. Conclusion: Our results reveal a novel mechanism, whereby caveolin-1 positively regulates human HCC cell invasiveness by coordinating VEGF-induced angiogenesis.

A TRAIL-Delivered Lipoprotein-Bioinspired Nanovector Engineering Stem Cell-Based Platform for Inhibition of Lung Metastasis of Melanoma.

Genetically engineered mesenchymal stem cells (MSCs), as non-viral gene delivery platforms, are rapidly evolving in tumor therapy due to their low immunogenicity and natural tumor-homing capacity. Methods: In this paper, we selected reconstituted high-density lipoprotein (rHDL), a lipoprotein-bioinspired nanovector with specific binding ability to scavenger receptor B type I (SR-BI) expressed on MSCs, as a transfection agent to genetically modify MSCs. pDNA encoding tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) was used as a functional gene to be transfected into the nucleus of MSCs for TRAIL expression. Lauric acid-coupled polyethyleneimine (PEI-LA) as an amphiphilic cationic polymer was synthesized to electrostatically bind to pDNA, and then incorporated into rHDL to form rHDL/PEI-LA/pDNA nanoparticles. Results: The nanoparticles exhibited homogenous particle size and excellent serum stability in vitro. Meanwhile, this SR-BI-targeted rHDL performed efficient intracellular gene delivery, specific lysosome-independent mechanism of cellular uptake and high transfection of pDNA towards MSCs. Moreover, high TRAIL expression in MSCs was detected after rHDL-mediated transfection. In vitro and in vivo results indicated that genetically engineered MSCs could accurately target to B16F10 cells, thereby producing significant apoptosis-inducing effect on aggressive melanoma. Conclusion: TRAIL-expressing MSCs engineered by rHDL nanovector was an efficient and hypotoxic method for stem cells-based pulmonary melanoma metastasis-targeting therapy.

Targeted next­generation sequencing identifies two novel COL2A1 gene mutations in Stickler syndrome with bilateral retinal detachment.

Stickler syndrome is a group of inherited connective tissue disorders characterized by distinctive facial and ocular abnormalities, hearing loss and early­onset arthritis. The aim of the present study was to investigate the genetic changes in two Chinese patients with Stickler syndrome, manifested as bilateral retinal detachment and peripheral retinal degeneration. Complete ophthalmic examinations, including best­corrected visual acuity, slit­lamp examination and fundus examination, were performed. Genomic DNA was extracted from leukocytes of the peripheral blood collected from the patients, their unaffected family members and 200 unrelated control subjects from the same population. Next­generation sequencing of established genes associated with ocular disease was performed. A heterozygous collagen type II α1 chain (COL2A1) mutation c.1310G>C (p.R437P) in exon 21 was identified in Family 1 and a heterozygous COL2A1 mutation c.2302­1G>A in intron 34 was identified in Family 2. The functional effects of the mutations were assessed by polymorphism phenotyping (PolyPhen) and sorting intolerant from tolerant (SIFT) analysis. The c.1310G>C mutation was predicted to damage protein structure and function, and the c.2302­1G>A mutation was predicted to result in a splicing defect. The findings of the current study expand the established mutation spectrum of COL2A1, and may facilitate genetic counseling and development of therapeutic strategies for patients with Stickler syndrome.

Intraoperative ultrasonography in laparoscopic partial nephrectomy for intrarenal tumors.

OBJECTIVE: To evaluate the feasibility and efficacy of intraoperative ultrasonography in laparoscopic partial nephrectomy (LPN) for intrarenal tumors. PATIENTS AND METHODS: All patients who underwent LPN for renal tumors in our institution from January 2010 to October 2016 were assessed retrospectively. Patients were divided into two groups, the first with totally intrarenal tumors (TIT group), defined as a solid renal mass with no exophytic element on both preoperative and intraoperative evaluations, and the second with exophytic tumors (control group). General information and perioperative data of the two groups were compared, including tumor characteristics, operative time, estimated blood loss, warm ischemia time and pathological findings. Intraoperative laparoscopic ultrasonography (ILUS) was used to precisely locate and delineate the TIT border, as well as seeking for other suspected lesions. RESULTS: We identified 583 patients who underwent LPN in our center, including 46 in the TIT and 537 in the control group. All patients in the TIT group were evaluated by ILUS, and all TIT procedures were successfully performed with only one conversion to open surgery. The mean tumor sizes in the TIT and control groups were 2.42 ± 0.46 cm and 3.29 ± 1.43 cm (p < 0.001), respectively. The TIT group's R.E.N.A.L. nephrometry score was higher than that of the control group (median 8.5 vs 6.0, p < 0.001), and their mean operation times were 127.2 ± 16.0 min and 120.1 ± 19.2 min, respectively. Mean estimated blood loss was higher in the TIT than in the control group (161.3 ml vs 136.6 ml, p = 0.003). Mean warm ischemia time differed in the TIT and control groups (22.2 ± 6.4 vs 20.6 ± 4.7 min, p = 0.105), but not significantly. Rates of open conversion and positive margins, as well as rates of major postoperative complications, pathological findings, and 1-month changes in renal function, were similar in the two groups. CONCLUSION: Intraoperative ultrasonography is technically feasible in patients undergoing LPN for TITs. This method may reduce the need for radical nephrectomy in patients with endogenic renal masses.

Regulatory B cells induced by ultraviolet B through toll-like receptor 4 signalling contribute to the suppression of contact hypersensitivity responses in mice.

BACKGROUND: Ultraviolet (UV) B irradiation is known to suppress contact hypersensitivity (CHS) responses in mouse models by suppressing immune responses. However, the cellular mechanisms responsible for UVB-induced systemic suppression remain unclear. Regulatory B cells have been reported to play an inhibitory role during CHS. It is presently unknown whether regulatory B cells contribute to the effect of UVB phototherapy. OBJECTIVE: To investigate the inductive effect of UVB on regulatory B cells and the underlying mechanisms by using a CHS mouse model. METHODS: CHS was induced with oxazolone, and evaluated by histopathology, flow cytometry, and quantitative real-time polymerase chain reaction. RESULT: We found that UVB irradiation induced regulatory B cell expansion and ameliorated CHS. UVB-induced regulatory B cells contribute to systemic immunosuppression by inhibiting the proliferation of T cells. Moreover, we determined that toll-like receptor (TLR) 4, the expression of which was upregulated in B cells after UVB exposure, played an essential role in the induction of regulatory B cells. CONCLUSION: Our data identified regulatory B cells as regulators of UVB-induced immunosuppression in CHS, and suggest the importance of the UVB-TLR4 axis in the generation of regulatory B cells.

Clinical and next-generation sequencing findings in a Chinese family exhibiting severe familial exudative vitreoretinopathy.

Familial exudative vitreoretinopathy (FEVR) is a rare hereditary retinal disorder characterized by the premature arrest of vascularization in the peripheral retina. The aim of the present study was to characterize the clinical presentations of a Chinese family affected by bilateral severe FEVR, and to identify the underlying genetic variations. One family that presented with bilateral FEVR was recruited for this study. Comprehensive ophthalmic examinations, including best­corrected visual acuity, slit­lamp examination, fundus photography, fundus fluorescein angiography imaging and electroretinogram were performed. Genomic DNA was extracted from leukocytes of the peripheral blood collected from the affected and unaffected family members, as well as 200 unrelated control subjects from the same population. Next­generation sequencing of the candidate genes associated with ocular diseases was performed, and the identified mutations were validated by conventional polymerase chain reaction­based sequencing. The functional effects of the mutations were analyzed by polymorphism phenotyping (PolyPhen) and sorting intolerant from tolerant (SIFT). One heterozygous ATP binding cassette subfamily A member 4 (ABCA4) c.5693G>A (p.R1898H) mutation in exon 40 and one heterozygous LDL receptor related protein 5 (LRP5) c.260T>G (p.I87S) mutation in exon 2 were identified in this family. To the best of our knowledge, the ABCA4 c.5693G>A (p.R1898H) mutation has not been reported in FEVR, and the LRP5 c.260T>G (p.I87S) mutation is a novel mutation. PolyPhen and SIFT predicted that the amino acid substitution R1898H in protein ABCA4 is benign, whereas the amino acid substitution I87S in protein LRP5 is damaging. A single nucleotide polymorphism c.266A>G (p.Q89R, rs41494349) was identified in exon 2 of LRP5. These findings expand the mutation spectrums of ABCA4 and LRP5, and will be valuable for genetic counseling and development of therapeutic interventions for patients with FEVR.

Two heterozygous mutations identified in one Chinese patient with bilateral macular coloboma.

Congenital macular coloboma is characterized by defined punched out atrophic lesions of the macula. The present study aimed to investigate the genetic alterations of one Chinese sporadic patient with bilateral large macular coloboma. Complete ophthalmic examinations, including best­corrected visual acuity, slit­lamp examination, fundus examination, fundus photograph and fundus fluorescein angiography imaging, Pentacam, and optical coherence tomography were performed on the patient. Genomic DNA was extracted from leukocytes in a peripheral blood sample collected from the patient, the patient's unaffected family members and from 200 unrelated control subjects from the same population. Next­generation sequencing of the known genes involved in ocular disease was performed. The functional effects of the mutation were analyzed using Polymorphism Phenotyping (PolyPhen) and Sorting Intolerant From Tolerant (SIFT). One heterozygous bestrophin 1 (BEST1) mutation c.1037C>A (p.Pro346His, p.P346H) in exon 9 and one heterozygous regulating synaptic membrane exocytosis 1 (RIMS1) mutation c.3481A>G (p.Arg1161Gly, p.R1161G) in exon 23 were identified in the patient being investigated, but not in the unaffected family members or unrelated control subjects. Polyphen and SIFT predicted that the amino acid substitution p.P346H in the BEST1 protein is damaging. In addition, Polyphen predicted that the amino acid substitution p.R1161G in the RIM1 protein is damaging. The results of the current study have increased the mutation spectrums of BEST1 and RIMS1, and are valuable for improving the current genetic counseling process and developing novel therapeutic interventions for patients with macular coloboma.