Extensive utilization increases the exposure of humans to Ag nanoparticles (NPs) via the oral pathway. To comprehensively address the action of Ag NPs to the gastrointestinal systems in real situations, i.e., the long-term low-dose exposure, we evaluated and compared the toxicity of three Ag NPs (20-30 nm with different surface coatings) to the human intestine cell Caco-2 after 1-day and 21-day exposures, using various biological assays. In both the short- and long-term exposures, the variety of surface coating predominated the toxicity of Ag NPs in a descending order of citrate-coated Ag NP (Ag-CIT), bare Ag NP (Ag-B), and poly (N-vinyl-2-pyrrolidone)-coated Ag NP (Ag-PVP). The short-term exposure induced cell growth inhibition and death. The cell viability loss appeared after cells were exposed to 0.7 μg/mL Ag-CIT, 0.9 μg/mL Ag-B or >1.0 μg/mL Ag-PVP for 24 h. The short-term and higher-dose exposure also induced reactive oxygen species (ROS) generation, mitochondrial damage, cell membrane leakage, apoptosis, and inflammation (IL-8 level). The long-term exposure only inhibited the cell proliferation. After 21-day exposure to 0.4 μg/mL Ag-CIT, the cell viability dropped to less than 50%, while cells exposed to 0.5 μg/mL Ag-PVP remained normal as the control. Generally, 0.3 μg/mL is the non-toxic dose for the long-term exposure of Caco-2 cells to Ag NPs in this study. However, cells presented inflammation after exposure to Ag NPs with the non-toxic dose in the long-term exposure.

Mucosal melanoma (MM) is the second most common melanoma subtype in Asian populations. Deregulation of microRNAs (miRNAs) has been extensively investigated in various cancers, including cutaneous melanoma. However, the roles of miRNAs in MM are unclear. In this study, we carried out miRNA profiling in MM, and we investigated the clinical and biological roles of miR-23a-3p in MM. Methods: miRNA expression in MM was profiled by miRNA microarray analysis. The expression of miR-23a-3p was quantitated by qRT-PCR in a cohort of 117 patients with MM, and its prognostic significance was evaluated. The biological effect of miR-23a-3p was demonstrated by both in vitro and in vivo studies through ectopic expression of miR-23a-3p. The target gene of miR-23a-3p and molecular pathway influenced by it was characterized using in silico target prediction tools, dual luciferase reporter assays, knockdown, and rescue experiments. Results: Microarray and qRT-PCR results showed that the miR-23a-3p level was substantially lower in MM, and low miR-23a-3p expression was significantly associated with poor outcomes. Ectopic expression of miR-23a-3p suppressed MM cell proliferation, migration, invasion, and tumorigenicity, indicating that miR-23a-3p has a tumor-suppressive role in MM. Mechanistic investigations identified adenylate cyclase 1 (ADCY1) as a direct target of miR-23a-3p in MM, and knockdown of ADCY1 recapitulated all the phenotypic characteristics of miR-23a-3p overexpression. Targeting of ADCY1 by miR-23a-3p resulted in the suppression of cyclic adenosine monophosphate (cAMP) and mitogen-activated protein kinase (MAPK) signaling pathways. Conclusions: Our data highlight the molecular etiology and clinical significance of miR-23a-3p in MM and reveal its major target and biological function. miR-23a-3p may represent a new prognostic biomarker or therapeutic target in MM.

Renal cell carcinoma (RCC) accounts for approximately 3% of adult malignancies, and the incidence of RCC continues to rise worldwide. Although RCC can be treated with surgery at an early stages, the five-year survival rates have been observed to decline dramatically in patients with advanced disease. Most patients with RCC treated with cytotoxic or targeted drugs will develop resistance at some point during therapy. Thus, it is necessary to identify novel therapeutic targets for RCC. Here, we found that RANBP2-type and C3HC4-type zinc finger-containing 1 (RBCK1) expression was upregulated in human RCC samples. Analysis of multiple public databases revealed the correlation between RBCK1 expression and poor prognosis in RCC patients. Subsequently, we performed RBCK1 depletion experiments in RCC cells that severely affected the in vivo and in vitro proliferation of renal cancer cells. The effects of RBCK1 on cell proliferation could be rescued with p53 expression knockdown in two cell lines expressing wild-type p53. Further experiments demonstrated that RBCK1 could facilitate p53 poly-ubiquitination and degradation by direct interaction with p53. Together, our results show that RBCK1 may serve as a promising target for RCC therapy by restoring p53 functions.

Lanthanide-doped upconversion nanoparticles can convert long wavelength excitation radiation to short wavelength emission. They have great potential in biomedical applications, such as bioimaging, biodetection, drug delivery, and theranostics. However, there is little information available on their bioavailability and biological effects after oral administration. In this study, we systematically investigated the bioavailability, biodistribution, and toxicity of silica-coated upconversion nanoparticles administrated by gavage. Our results demonstrate that these nanoparticles can permeate intestinal barrier and enter blood circulation by microstructure observation of Peyer's patch in the intestine. Comparing the bioavailability and the biodistribution of silica-coated upconversion nanoparticles with oral and intravenous administration routes, we found that the bioavailability and biodistribution are particularly dependent on the administration routes. After consecutive gavage for 14 days, the body weight, pathology, Zn and Cu level, serum biochemical analysis, oxidative stress, and inflammatory cytokines were studied to further evaluate the potential toxicity of the silica-coated upconversion nanoparticles. The results suggest that these nanoparticles do not show overt toxicity in mice even at a high dose of 100 mg/kg body weight.

Breast cancer is the leading cause of cancer death in women. Chemotherapy to inhibit the proliferation of cancer cells is considered to be the most important therapeutic strategy. The development of long-circulating PEG and targeting liposomes is a major advance in drug delivery. However, the techniques used in liposome preparation mainly involve conventional liposomes, which have a short half-life, high concentrations in the liver and spleen reticuloendothelial system, and no active targeting.

Rationale: Tumor energy metabolism has been a well-appreciated target of cancer therapy; however, the metabolism change of cancer cells between oxidative phosphorylation and glycolysis poses a challenge to the above. In this study, we constructed an innovative mitochondrion-targeted supramolecular "nano-boat" based on peptide self-assembly for tumor combined chemo-radiotherapy by simultaneously inhibiting the dual energy metabolism. Methods: A lipophilic self-assembled peptide and a positively charged cyclen were integrated to fabricate a brand new mitochondrion-targeted nano-platform for the first time. The indices of mitochondrial dysfunction including mitochondrial membrane potential, apoptosis proteins expression and ultrastructure change were evaluated using a JC-1 probe, western blotting and biological transmission electron microscopy, respectively. Energy metabolism assays were conducted on a Seahorse XF24 system by detecting the oxygen consumption rate and the glycolytic proton efflux rate. The radio-sensitization effect was investigated by colony formation, the comet assay, and γ-H2AX staining. Results: The supramolecular "nano-boat" could selectively kill cancer cells by much higher enrichment and reactive oxygen species generation than those in normal cells. In the cancer cells treated with the supramolecular "nano-boat", the dysfunctional morphological changes of the mitochondrial ultrastructure including swelling and pyknosis were evidently observed, and the endogenous mitochondrial apoptosis pathway was effectively triggered by abundant of cytochrome C leaking out. Concurrently, the dual metabolic pathways of glycolysis and oxidative phosphorylation were severely inhibited. More importantly, the supramolecular "nano-boat" displayed an excellent radio-sensitization effect with a sensitization enhancement ratio value as high as 2.58, and hence, in vivo efficiently combining radiotherapy yielded an enhanced chemo-radiotherapy effect. Conclusion: Our study demonstrated that the rationally designed peptide-based "nano-boat" could efficiently induce cancer cell apoptosis by the energy metabolism inhibition involving multiple pathways, which may provide the motivation for designing novel and universal mitochondria-targeted drug delivery systems for cancer therapy.

Chemotherapy is still the main first-line treatment for advanced metastatic gastric cancer, but it has the limitations of serious side effects and drug resistance. Conventional liposome has been substantially used as drug carriers, but they lack targeting character with lower drug bioavailability in tumor tissues. Based on the above problems, a novel estrogen-targeted PEGylated liposome loaded with oxaliplatin (ES-SSL-OXA) was prepared to further improve the metabolic behavior, the safety profile, and the anti-tumor efficacy of oxaliplatin.

Patients with intrahepatic cholangiocarcinoma (ICC) require chemotherapy due to late detection, rapid disease progression, and low surgical resection rate. Tumor cell lines are extremely important in cancer research for drug discovery and development. Here, we established and characterized a new intrahepatic cholangiocarcinoma cell line, ICC-X1. STR testing confirmed the absence of cross-contamination and high similarity to the original tissue. ICC-X1 exhibited typical epithelial morphology and formed tumor spheres in the suspension culture. The population doubling time was approximately 48 h. The cell line had a complex hypotriploid karyotype. The cell line exhibited a strong migration ability in vitro and cell inoculation into BALB/c nude mice led to the formation of xenografts. Additionally, ICC-X1 cells were sensitive to gemcitabine and paclitaxel but resistant to 5-fluorouracil and oxaliplatin. RNA sequencing revealed that the upregulated cancer-related genes were mainly enriched in several signaling pathways, including the TNF signaling pathway, NOD-like receptor signaling pathway, and NF-κB signaling pathway. The downregulated cancer-related genes were mainly enriched in the Rap1 signaling pathway and Hippo signaling pathway among other pathways. In conclusion, we have created a new ICC cell line derived from Chinese patients. This cell line can be used as a preclinical model to study ICC, specifically tumor metastasis and drug resistance mechanisms.

Although nasal obstruction (NO) during growth causes maxillofacial growth suppression, it remains unclear whether eliminating the NO affects maxillary and mandibular growth differentially. We aimed to clarify whether eliminating NO can help regain normal maxillofacial growth and to determine the optimal intervention timing. Forty-two 4-week-old male Wistar rats were randomly divided into six groups. Their left nostril was sutured to simulate NO over different durations in the experimental groups; the sutures were later removed to resume nasal breathing. Maxillofacial morphology was assessed using microcomputed tomography. Immunohistochemical changes in hypoxia-inducible factor (HIF)-1α, osteoprotegerin (OPG), and receptor activator of nuclear factor kappa-B ligand (RANKL) of the condylar cartilage were evaluated to reveal the underlying mechanisms of these changes. Maxillary length was significantly lower in rats with NO for ≥5 weeks. In groups with NO for ≥7 weeks, the posterior mandibular length, ramus height, thickness of the hypertrophic cell layer in the condylar cartilage, HIF-1α levels, and RANKL levels were significantly lower and OPG levels and RANKL/OPG were significantly higher than those in the control group. Our findings suggest that eliminating NO is effective in regaining maxillofacial growth. Moreover, the optimal timing of intervention differed between the maxilla and mandible.

Tripterygium glycoside tablet (TGT), as a common clinical drug, can easily cause liver damage due to the narrow therapeutic window. Glycyrrhizic acid (GA) has a hepatoprotective effect, but the characteristics and mechanism of GA's impact on TGT-induced acute liver injury by regulating oxidative stress remain unelucidated. In this study, TGT-induced acute liver injury models were established in vitro and in vivo. The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (AKP), superoxide dismutase (SOD), malondialdehyde (MDA), glutathione (GSH), catalase (CAT), lactate dehydrogenase (LDH), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6) were quantified. The anti-apoptotic effect of GA was tested using flow cytometry. Potential target proteins of GA were profiled via activity-based protein profiling (ABPP) using a cysteine-specific (IAA-yne) probe. The results demonstrate that GA markedly decreased the concentrations of ALT, AST, AKP, MDA, LDH, TNF-α, IL-1β and IL-6, whereas those of SOD, GSH and CAT increased. GA could inhibit TGT-induced apoptosis in BRL-3A cells. GA bound directly to the cysteine residue of PKM2. The CETSA and enzyme activity results validate the specific targets identified. GA could mitigate TGT-induced acute liver injury by mediating PKM2, reducing oxidative stress and inflammation and reducing hepatocyte apoptosis.

The aim of this study was to investigate the changes in bone marrow fatty acids early after ovariectomy-induced osteoporosis in rats, and explore the potential function of the bone marrow fatty acids. Ninety-six female Sprague Dawley rats (12 weeks) were randomly divided into an ovariectomized (OVX) group and Sham group (N = 48/group) and received ovariectomy or Sham surgery, respectively. After 3, 5, 7,14, 21 and 28 days, eight rats in each group were sacrificed to detect the composition of bone marrow fatty acids by means of gas chromatography-mass spectrometry and evaluate the trabecular bone microarchitecture by means of microCT. Bone marrow rinsing fluid and serum were collected for the detection of nitric oxide synthase/nitric oxide (NOS/NO) and bone metabolism related parameters, respectively. Our results demonstrated that the bone microstructure was damaged significantly from 14 days after OVX surgery onwards. Sample clustering and group separation were observed between the OVX group and Sham group 3 and 14 days after surgery, which suggested the role of bone marrow fatty acids in the early stage of postmenopausal osteoporosis. Palmitoleate, myristate and arachidonate were found to play an important role in classification between the OVX group and Sham group on the 3rd day after surgery (VIP > 1, p < 0.05). Palmitoleate, myristate, alpha linolenate, stearate and eicosenoate were found to play an important role in classification between the OVX group and Sham group on the 14th day after surgery (VIP > 1, p < 0.05). The levels of myristate, palmitoleate, alpha linolenate and eicosenoate were significantly decreased in the OVX group, while the levels of arachidonate and stearate were significantly increased in OVX group (p < 0.05). Additionally, myristate, palmitoleate, alpha linoleate and eicosenoate were negatively correlated with C-terminal telopeptide of type 1 collagen (CTX-1, a bone resorption marker), while arachidonate was negative correlated with osteocalcin (OCN, a bone formation marker) (p < 0.05). A significant correlation was also found between eicosenoate and NOS (p < 0.05). Profound bone marrow fatty acids changes have taken place in the early stage of post-menopausal osteoporosis. They may affect bone formation though affecting the differentiation and function of osteoclasts or osteoblasts, respectively. The NOS/NO system may mediate the influence of eicosenoate on bone formation.

Mucosal melanoma with poor prognosis is a common histopathologic subtype of melanoma among Chinese and other Asian peoples. Regulated microRNAs (miRNAs) have been reported as oncogenes or tumour suppressors in melanoma. However, the roles of specific miRNAs in mucosal melanoma remain largely unknown. Here, we aimed to assess the biological functions, molecular mechanisms and clinical potential of miR-let-7b and miR-let-7c in mucosal melanoma.

Ovarian cancer is the most lethal gynecologic malignancy. The combination of paclitaxel (PTX) and carboplatin (CBP) is the first-line remedy for clinical ovarian cancer. However, due to the limitations of adverse reaction and lacking of targeting ability, the chemotherapy of ovarian cancer is still poorly effective. Here, a novel estrone (ES)-conjugated PEGylated liposome co-loaded PTX and CBP (ES-PEG-Lip-PTX/CBP) was designed for overcoming the above disadvantages.

Immune modulation has been recognized as an effective anti-osteoporosis strategy since the pivotal role of the RANK/RANKL/OPG signaling in bone metabolism and remodeling was discovered. To investigate the potential preventive and/or therapeutic effects of immune modulator protein Ling Zhi-8 (LZ-8) on osteoporosis, the osteoporosis animal model was established in Wistar rats by intramuscular injection of dexamethasone (DEX), namely glucocorticoids-induced osteoporosis (GIOP) rat model. To investigate the potential preventive effect of rLZ-8 on GIOP, we co-treated the rats with DEX and rLZ-8 intraperitoneally during the GIOP modeling stage and analyze the bone mass measured by bone mineral content (BMC) and bone mineral density (BMD), as well as levels of phosphorus (Pi), calcium (Ca2+) and hydroxyproline (HOP) in femur of GIOP rats. Consistently, all results suggested that rLZ-8 could prevent bone loss in the femurs of GIOP rats. Through analyzing the trabeculae morphology and the trabeculae amount by H&E staining, we found rLZ-8 could also improve the structural deterioration in femurs of GIOP rats. In order to further verify the results and its mechanism obtained from bone analysis, multiple biomarkers, including minerals metabolism (Pi and Ca2+), bone formation markers (osteocalcin, ALP and IGF-1), bone resorption markers (TRACP5b, CTX-1 and HOP), cytokines (IL-1β, IL-6 and TNF-α), oxidative stress indicators (GSH-px, SOD and MDA) and hormone molecules (testosterone, estradiol, calcitonin and parathyroid hormone) have been detected in serum or urine of rats. Results of these biomarkers in serum or urine confirmed rLZ-8's protective effect in GIOP. Through analyzing the relative expression level of OPG and RANKL in femurs via western blot, we foundrLZ-8 could increase OPG/RANKL ratio which could impede osteoclastogenesis process. To test the potential therapeutic effect of rLZ-8 on successfully generated GIOP rats, we administrated rLZ-8 to rats for three weeks starting from the ending day of 7 weeks treatment of DEX. We found rLZ-8 could also reverse the bone loss in GIOP rats. Through the BWs and organ coefficient analysis, we found rLZ-8 has little toxicity to the rats. Our results suggested that rLZ-8 may be developed into promising anti-osteoporosis drug with both preventive and therapeutic properties.

Coexistence of enhancer of zeste homolog 2 (EZH2) and BRAF gene aberrations has been described in many cancer types. In this study, we aim to explore the coexistence status of BRAF V600E mutation and the copy number variation of EZH2 and explore the potential of this combination as a therapeutic target.

Aristolochic acid (AA), mainly derived from herbal Aristolochia and Asarum plants, was listed as a human carcinogen class I in 2002. Aristolochic acid nephropathy (AAN) is a rapidly progressive tubulointerstitial nephritis and urothelial cancer caused by AA. However, the targeting molecular mechanisms of AAs-induced nephrotoxicity are largely unclear. This study aims to dissect targeting molecular mechanisms of AA-induced nephrotoxicity. Activity-based protein profiling (ABPP) in combination with cellular thermal shift assay (CETSA) was performed to identify the AAs binding target proteins. Our data indicated that several key enzymes in the metabolic process and mitochondrial respiration including IDH2 and MDH2 (Krebs cycle), PKM and LDH (aerobic respiration), FASN (fatty acid beta-oxidation), HK2 (glucose metabolism), and ATP synthase were identified as directly binding targets of AAs. Metabolomics and oxygen consumption rate (OCR) experiments further confirmed that AAs targeting proteins disrupted metabolic biosynthesis processes and impaired mitochondrial functions. Ultimately, AAs induced renal cells apoptosis by disturbing various biological processes. Cumulatively, AAs may directly bind to key proteins involved in the metabolic process and mitochondrial homeostasis, and finally induce aristolochic acid nephropathy. Our findings provide novel insight into underlying mechanisms of AAs-induced kidney toxicity, which may help to develop therapeutic strategies for AAN.

The clinical outcome of resectable non-small-cell lung cancer (NSCLC) patients receiving neoadjuvant chemoimmunotherapy is good but varies greatly. In addition, the pathological response after neoadjuvant chemoimmunotherapy is significantly associated with survival outcomes. The aim of this retrospective study was to identify which population of patients with locally advanced and oligometastatic NSCLC has a favorable pathological response after neoadjuvant chemoimmunotherapy. NSCLC patients treated with neoadjuvant chemoimmunotherapy were enrolled between February 2018 and April 2022. Data on clinicopathological features were collected and evaluated. Multiplex immunofluorescence was performed on pre-treatment puncture specimens and surgically resected specimens. In total, 29 patients with stages III and IV locally advanced or oligometastatic NSCLC who received neoadjuvant chemoimmunotherapy and R0 resection were enrolled. The results showed that 55% (16/29) of patients had a major pathological response (MPR) and 41% (12/29) of patients had a complete pathological response (pCR). In the stroma area of the pre-treatment specimen, the higher infiltration of CD3+ PD-L1+ tumor-infiltrating lymphocytes (TILs) and the lower infiltration of CD4+ and CD4+ FOXP3+ TILs were more likely to appear in patients with pCR. However, in the tumor area, the higher infiltration of CD8+ TILs was more likely to appear in patients with non-MPR. In the post-treatment specimen, we found increased infiltration of CD3+ CD8+ , CD8+ GZMB+ , and CD8+ CD69+ TILs and decreased infiltration of PD-1+ TILs both in the stroma and tumor areas. Neoadjuvant chemoimmunotherapy achieved an MPR rate of 55% and induced greater immune infiltration. In addition, we observed that the baseline TILs and their spatial distribution correlate to the pathological response.

Hepatic stellate cells (HSCs) are essential drivers of fibrogenesis. Inducing activated-HSC apoptosis is a promising strategy for treating hepatic fibrosis. 18beta-glycyrrhetinic acid (18β-GA) is a natural compound that exists widely in herbal medicines, such as Glycyrrhiza uralensis Fisch, which is used for treating multiple liver diseases, especially in Asia. In the present study, we demonstrated that 18β-GA decreased hepatic fibrosis by inducing the apoptosis in activated HSCs. 18β-GA inhibited the expression of α-smooth muscle actin and collagen type I alpha-1. Using a chemoproteomic approach derived from activity-based protein profiling, together with cellular thermal shift assay and surface plasmon resonance, we found that 18β-GA covalently targeted peroxiredoxin 1 (PRDX1) and peroxiredoxin 2 (PRDX2) proteins via binding to active cysteine residues and thereby inhibited their enzymatic activities. 18β-GA induced the elevation of reactive oxygen species (ROS), resulting in the apoptosis of activated HSCs. PRDX1 knockdown also led to ROS-mediated apoptosis in activated HSCs. Collectively, our findings revealed the target proteins and molecular mechanisms of 18β-GA in ameliorating hepatic fibrosis, highlighting the future development of 18β-GA as a novel therapeutic drug for hepatic fibrosis.

The purpose of this study was to optimize the preparation method of injectable Octreotide microspheres. To explore the correlation between the solvent system and the general properties of microspheres to reduce burst release and enable them to be used for portal hypertension. Octreotide microspheres were prepared by modified double emulsion solution evaporation method after optimizing preparation conditions. The results showed that Octreotide microspheres had a particle size of 57.48 ± 15.24 μm, and the initial release was significantly reduced. In vitro release and in vivo pharmacokinetic data indicated that Octreotide was released stably within 1200 h. The effects on portal vein pressure, liver tissue morphology and other related indexes were observed after administration. As obvious results, injection of Octreotide microspheres could significantly reduce portal vein pressure and reduce the portal vein lumen area in experimental cirrhotic portal hypertensive rats. The optimized Octreotide PLGA microsphere preparation has been proved to have a good effect on PHT in vivo after detecting aminotransferase (AST) and alanine aminotransferase (ALT) activity, liver tissue hydroxyproline (Hyp) content, serum and liver tissue malondialdehyde (MDA) levels, plasma prostacyclin (PGI2) levels, and liver tissue tumor necrosis factor (TNFα) content. In addition, serum and liver tissue superoxide dismutase (SOD) activity and liver tissue glutathione (GSH) content, plasma thromboxane (TXA2), serum nitric oxide (NO), liver tissue nitric oxide synthase (NOS), and plasma and liver tissue endothelin (ET) were significantly increased.

G2 and S-phase expressed 1 (GTSE1) regulates cell cycle progression in human cancers. However, its significance and mechanism of action in acral melanoma (AM) remain unknown. In the present study, we found that GTSE1 expression was upregulated in advanced stage/metastatic AM tissues and metastatic cell lines, and correlated with higher stage (P = .028) and poor disease-free survival (DFS) in patients with AM (P = .003). Cox regression assays validated GTSE1 expression to be an independent prognostic factor of DFS for patients with AM (P = .004). Ectopic expression of GTSE1 enhanced primary AM cell proliferation, invasion, and migration. Loss-of-function in GTSE1 attenuated metastatic AM cell proliferation and metastatic ability in vitro and in vivo. We additionally observed that inhibition of migration and invasion occurred concomitantly with a GTSE1 knockdown-mediated increase in E-cadherin and decreases in N-cadherin and Slug. We further showed that integrin subunit alpha 2 (ITGA2) interacts with GTSE1 and is a downstream effector of GTSE1. Further, ITGA2 levels were positively correlated with GTSE1 expression in human AM tissues. Ectopic ITGA2 expression rescued siGTSE1-mediated inhibition of migration and invasion, thereby restoring epithelial-to-mesenchymal transition (EMT). In conclusion, GTSE1 expression promotes AM progression and correlates with clinical outcomes of patients with AM, and may represent a promising therapeutic target to suppress AM progression.