PTEN Deficiency Facilitates Exosome Secretion and Metastasis in Cholangiocarcinoma by Impairing TFEB-mediated Lysosome Biogenesis.

BACKGROUND & AIMS: In eukaryotes, the ubiquitin-proteasome system and the autophagy-lysosome pathway are essential for maintaining cellular proteostasis and associated with cancer progression. Our previous studies have demonstrated that phosphatase and tensin homolog (PTEN), one of the most frequently mutated genes in human cancers, limits proteasome abundance and determines chemosensitivity to proteasome inhibitors in cholangiocarcinoma (CCA). However, whether PTEN regulates the lysosome pathway remains unclear. METHODS: We tested the effects of PTEN on lysosome biogenesis and exosome secretion using loss- and gain-of-function strategies in CCA cell lines. Using in vitro dephosphorylation assays, we explored the regulatory mechanism between PTEN and the key regulator of lysosome biogenesis, transcription factor EB (TFEB). Using the migration assays, invasion assays, and trans-splenic liver metastasis mouse models, we evaluated the function of PTEN deficiency, TFEB-mediated lysosome biogenesis, and exosome secretion on tumor metastasis. Moreover, we investigated the clinical significance of PTEN expression and exosome secretion by retrospective analysis. RESULTS: PTEN facilitated lysosome biogenesis and acidification through its protein phosphatase activity to dephosphorylate TFEB at Ser211. Notably, PTEN deficiency increased exosome secretion by reducing lysosome-mediated degradation of multi-vesicular bodies, which further facilitated the proliferation and invasion of CCA. TFEB agonist curcumin analog C1 restrained the metastatic phenotype caused by PTEN deficiency in mouse models, and we highlighted the correlation between PTEN deficiency and exosome secretion in clinical cohorts. CONCLUSIONS: In CCA, PTEN deficiency impairs lysosome biogenesis to facilitate exosome secretion and cancer metastasis in a TFEB phosphorylation-dependent manner.

A 7-Day Decitabine-Included Conditioning Regimen Accelerated Donor Hematopoietic Engraftment while Reduced the Occurrence of Mucositis without Interfering with Prognosis.

INTRODUCTION: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the standard and curative treatment strategy for patients with hematologic malignancies. Recently, decitabine-included regimens have been investigated by several studies including ours, which may prevent relapse of primary malignant diseases. METHODS: This study was to retrospectively evaluate a 7-day decitabine-included regimen with reduced dose of idarubicin for patients with hematologic malignancies who underwent allo-HSCT. RESULTS: A total of 84 patients were enrolled, including 24 cases in 7-day and 60 cases in 5-day decitabine groups, respectively. Patients conditioned with 7-day decitabine regimen showed accelerated neutrophil (12.05 ± 1.97 vs. 13.86 ± 3.15; u = 9.309, p < 0.001) and platelet (16.32 ± 6.27 vs. 21.37 ± 8.57; u = 8.887, p < 0.001) engraftment compared with those treated with 5-day decitabine regimen. Patients in the 7-day decitabine group showed a significantly lower incidence rate of total (50.00% [12/24] versus 78.33% [47/60]; χ2 = 6.583, p = 0.010) and grade III or above (4.17% [1/24] vs. 31.67% [19/60]; χ2 = 7.147, p = 0.008) oral mucositis compared to those in the 5-day decitabine group. However, the occurrence of other major complications post-allo-HSCT and outcomes of patients in these two groups were comparable. CONCLUSION: These results demonstrate that this 7-day decitabine-contained new conditioning regimen seems to be feasible and safe for patients with myeloid neoplasms who receive allo-HSCT, and a large-scale prospective study is needed to confirm the findings of this study.

[Effects of total ginsenosides from Panax ginseng stems and leaves on gut microbiota and short-chain fatty acids metabolism in acute lung injury mice].

This study aimed to investigate the biological effects and underlying mechanisms of the total ginsenosides from Panax ginseng stems and leaves on lipopolysaccharide(LPS)-induced acute lung injury(ALI) in mice. Sixty male C57BL/6J mice were randomly divided into a control group, a model group, the total ginsenosides from P. ginseng stems and leaves normal administration group(61.65 mg·kg~(-1)), and low-, medium-, and high-dose total ginsenosides from P. ginseng stems and leaves groups(15.412 5, 30.825, and 61.65 mg·kg~(-1)). Mice were administered for seven continuous days before modeling. Twenty-four hours after modeling, mice were sacrificed to obtain lung tissues and calculate lung wet/dry ratio. The number of inflammatory cells in bronchoalveolar lavage fluid(BALF) was detected. The levels of interleukin-1ß(IL-1ß), interleukin-6(IL-6), and tumor necrosis factor-α(TNF-α) in BALF were detected. The mRNA expression levels of IL-1ß, IL-6, and TNF-α, and the levels of myeloperoxidase(MPO), glutathione peroxidase(GSH-Px), superoxide dismutase(SOD), and malondialdehyde(MDA) in lung tissues were determined. Hematoxylin-eosin(HE) staining was used to observe the pathological changes in lung tissues. The gut microbiota was detected by 16S rRNA sequencing, and gas chromatography-mass spectrometry(GC-MS) was applied to detect the content of short-chain fatty acids(SCFAs) in se-rum. The results showed that the total ginsenosides from P. ginseng stems and leaves could reduce lung index, lung wet/dry ratio, and lung damage in LPS-induced ALI mice, decrease the number of inflammatory cells and levels of inflammatory factors in BALF, inhibit the mRNA expression levels of inflammatory factors and levels of MPO and MDA in lung tissues, and potentiate the activity of GSH-Px and SOD in lung tissues. Furthermore, they could also reverse the gut microbiota disorder, restore the diversity of gut microbiota, increase the relative abundance of Lachnospiraceae and Muribaculaceae, decrease the relative abundance of Prevotellaceae, and enhance the content of SCFAs(acetic acid, propionic acid, and butyric acid) in serum. This study suggested that the total ginsenosides from P. ginseng stems and leaves could improve lung edema, inflammatory response, and oxidative stress in ALI mice by regulating gut microbiota and SCFAs metabolism.

A pilot study of viscoelastic agent to prevent recurrent vitreous hemorrhage after vitrectomy for proliferative diabetic retinopathy.

BACKGROUND: To evaluate the possibilty of preventing recurrent vitreous hemorrhage (RVH) after vitrectomy in proliferative diabetic retinopathy (PDR) patients with unabsorbed vitreous hemorrhage (VH) by intravitreal injection of viscoelastic agent (VA) at the end of the surgery and compared its effect with triamcinolone acetonide (TA). METHODS: This was a pilot prospective, observational study. PDR patients with VH who underwent vitrectomy were assigned to 3 groups according to the tamponade applicated at the end of the surgery, including VA group (intravitreally injected 1 ml VA if the retina was prone to bleed during the operation), TA group (intravitreally injected 2 mg TA when there was much exudates), or balanced salt solution (BSS) group (no tamponade). Then postoperative follow-up was performed routinely until 6 months after surgery. The primary outcome was the incidence of RVH, secondary outcome were the best-corrected visual acuity (BCVA) and introcular pressure (IOP). Cataract formation and other complication were also assessed. RESULTS: A total of 68 eyes, from 68 patients, were included. 18,18,32 eyes were enrolled in the VA group, TA group and BSS group, respectively. The integral incidence of RVH after vitrectomy was 5.6%, 5.6% and 12.5% respectively (P = 0.602). There was no early RVH in VA or TA group, whereas 3 early RVHs were identified in BSS group, however there was no significant difference (P = 0.171). Every group had one late RVH case. In all groups, final BCVA showed significant improvement compared to baseline. BCVA at any postoperative visit showed no significant differences among 3 groups. Mean IOP was higher 1 week after surgery in VA group compared with the other groups; however, in other times the differences were not significant. No cataract formation and other complication was noted in 3 groups. CONCLUSION: Intravitreal injection of VA or TA at the end of vitrectomy for PDR patients with unabsorbed VH tend to reduce the incidence of early RVH after vitrectomy similarly. As VA was preferred to applicate in the eyes that were prone to bleed, intravitreal injection of VA at the end of vitrectomy might be a promising method for preventing RVH in PDR patients.

Targeting hexokinase 2 increases the sensitivity of oxaliplatin by Twist1 in colorectal cancer.

Colorectal cancer (CRC) is the third most malignant tumour worldwide, with high mortality and recurrence. Chemoresistance is one of the main factors leading to metastasis and poor prognosis in advanced CRC patients. By analysing the Gene Expression Omnibus data set, we found higher hexokinase 2 (HK2) expression levels in patients with metastatic CRC than in those with primary CRC. Moreover, we observed higher enrichment in oxaliplatin resistance-related gene sets in metastatic CRC than in primary CRC. However, the underlying relationship has not yet been elucidated. In our study, HK2 expression was significantly elevated in CRC patients. Gene set enrichment analysis (GSEA) revealed multi-drug resistance and epithelial-mesenchymal transition (EMT) pathways related to high HK2 expression. Our results showed that knockdown of HK2 significantly inhibited vimentin and Twist1 expression and promoted TJP1 and E-cadherin expression in CRC cells. Additionally, transcriptional and enzymatic inhibition of HK2 by 3-bromopyruvate (3-bp) impaired oxaliplatin resistance in vitro and in vivo. Mechanistically, HK2 interacts with and stabilized Twist1 by preventing its ubiquitin-mediated degradation, which is related to oxaliplatin resistance, in CRC cells. Overexpression of Twist1 reduced the apoptosis rate by HK2 knockdown in CRC cells. Collectively, we discovered that HK2 is a crucial regulator that mediates oxaliplatin resistance through Twist1. These findings identify HK2 and Twist1 as promising drug targets for CRC chemoresistance.

RPB5-Mediating Protein Promotes Cholangiocarcinoma Tumorigenesis and Drug Resistance by Competing With NRF2 for KEAP1 Binding.

BACKGROUND AND AIMS: Cancer cell survival depends on the balance between reactive oxygen species production and scavenging, which is regulated primarily by NRF2 during tumorigenesis. Here, we demonstrate that deletion of RBP5-mediating protein (RMP) in an autonomous mouse model of intrahepatic cholangiocarcinoma (ICC) delays tumor progression. APPROACH AND RESULTS: RMP-overexpressing tumor cells exhibited enhanced tolerance to oxidative stress and apoptosis. Mechanistically, RMP competes with NRF2 for binding to the Kelch domain of KEAP1 (Kelch-like ECH-associated protein 1) through the E**E motif, leading to decreased NRF2 degradation via ubiquitination, thus increasing NRF2 nuclear translocation and downstream transactivation of antioxidant genes. This RMP-KEAP1-NRF2 axis promotes ICC tumorigenesis, metastasis, and drug resistance. Consistent with these findings, the RMP level in human ICC is positively correlated with the protein level of NRF2 and is associated with poor prognosis. CONCLUSION: These findings reveal that RMP is involved in the oxidative stress defense program and could be exploited for targeted cancer therapies.

Ultrastructural study of closed macular hole- preliminary application of a novel high magnification module combining with OCT.

BACKGROUND: As a novel high magnification module (HMM) combining with OCT (OCT-HMM) is able to detect the microstructure of retina, we apply it to explore the ultrastructure of the macula after closure of the idiopathic macular hole (IMH) by surgery. METHODS: This is an observational case series study in which patients with full-thickness IMHs who had undergone successful macular closure by vitrectomy and internal limiting membrane peeling and healthy subjects were recruited. After comprehensive ophthalmic examinations, the images of macular area were obtained and collected by professional operators using OCT-HMM. Then images were independently analyzed by 4 masked vitreoretinal specialists. RESULTS: A total of 24 IMH eyes and 42 healthy eyes were examined. HMM images were obtained in 10 IMH eyes. Among them, 4 eyes whose macula closed completely with recovery of photoreceptor layer presented a dark arc nasal to the fovea, oriented to the optic, and the notch of arc faced temporally. Six eyes in which the macula closed incompletely with photoreceptor cells loss revealed a dark ring with uneven bright spots inside. The other 14 eyes failed to obtain clear images by OCT-HMM. The contra lateral eyes of the patients and the healthy subjects' eyes succeeded to obtain the HMM images which displayed evenly grey background thickly covered with tiny bright dots that was in similar size and evenly and widely distributed and there no dark arc or ring. OCT B-scan and IR images could be acquired in all of the IMH and healthy eyes. CONCLUSION: The preliminary application of HMM has supplied us a brand-new insight into the microstructure of closed IMH. A dark arc sign could be detected with OCT-HMM in the macula which was functionally closed after surgery that was probably the healing mark on a microstructure photoreceptors level. Its existence and shape indicated that the functional closure followed by a retinal displacement mainly horizontally from temporal side to nasal side but not symmetric centripetally.

[Identification of key enzyme genes involved in biosynthesis of steroidal saponins and analysis of biosynthesis pathway in Polygonatum cyrtonema].

Steroidal saponins, which are the characteristic and main active constituents of Polygonatum, exhibit a broad range of pharmacological functions, such as regulating blood sugar, preventing cardiovascular and cerebrovascular diseases and anti-tumor. In this study, we performed RNA sequencing(RNA-Seq) analysis for the flowers, leaves, roots, and rhizomes of Polygonatum cyrtonema using the BGISEQ-500 platform to understand the biosynthesis pathway of steroidal saponins and study their key enzyme genes. The assembly of transcripts for four tissues generated 129 989 unigenes, of which 88 958 were mapped to several public databases for functional annotation, 22 813 unigenes were assigned to 53 subcategories and 64 877 unigenes were annotated to 136 pathways in KEGG database. Furthermore, 502 unigenes involved in the biosynthesis pathway of steroidal saponins were identified, of which 97 unigenes encoding 12 key enzymes. Cycloartenol synthase, the first key enzyme in the pathway of phytosterol biosynthesis, showed conserved catalytic domain and substrate binding domain based on sequence analysis and homology modeling. Differentially expressed genes(DEGs) were identified in rhizomes as compared to other tissues(flowers, leaves or roots).The 2 437 unigenes annotated by KEGG showed rhizome-specific expression, of which 35 unigenes involved in the biosynthesis of steroidal saponins. Our results greatly extend the public transcriptome dataset of Polygonatum and provide valuable information for the identification of candidate genes involved in the biosynthesis of steroidal saponins and other important secondary metabolites.

Six2 is negatively correlated with prognosis and facilitates epithelial-mesenchymal transition via TGF-ß/Smad signal pathway in hepatocellular carcinoma.

BACKGROUND: Increasing evidence indicates that Six2 contributes to tumorigenesis in various tumor including hepatocellular carcinoma (HCC). This study aimed to determine the role of Six2 in HCC and to elucidate the association of Six2 with clinical pathological characteristics. METHODS: The expressions of Six2 in HCC tumor, para-tumor tissue and portal vein tumor thrombus (PVTT) were detected by tissue microarray technique, immunohistochemistry, real-time RT-PCR and Western blotting. Chi-square and Kaplan-Meier analysis were used to analyze the correlation between Six2 expression and prognosis of HCC patients. Lentivirus mediated Six2 knockdown, spheroid formation assay, proliferation assay and subcutaneous tumor implantation were performed to determine the function of Six2. RESULTS: In 274 HCC samples, Six2 was strongly expressed. Kaplan-Meier analysis revealed that high expression of Six2 was correlated with a shorter overall survival (OS) and disease-free survival (DFS). Moreover, Six2 expression was associated with sex, alpha-fetoprotein, tumor size and portal vein invasion. Six2 was highly expressed in PVTT. Six2 knockdown inhibited HCC cell lines proliferation, migration, and self-renewal in vitro and in vivo. In addition, low-expression of Six2 weakened TGF-ß induced Smad4 activation and epithelial-mesenchymal transition in HCC cell lines. CONCLUSIONS: Elevated Six2 expression in HCC tumor patients was associated with negative prognosis. Upregulated Six2 promoted tumor growth and facilitated HCC metastasis via TGF-ß/Smad signal pathway.

[Transcriptome analysis of venom gland and identification of functional genes for snake venom protein in Agkistrodon acutus].

Agkistrodon acutus is a traditional Chinese herb medicine which has immunological regulation,anti-tumor,anti-inflammatory and analgesic effects,which is mainly used for the treatment of rheumatoid arthritis,ankylosing spondylitis,sjogren's syndrome and tumors. In order to excavate more important functional genes from A. acutus,the transcriptome of the venom gland was sequenced by the Illumina Hi Seq 4000,and 32 862 unigenes were assembled. Among them,26 589 unigenes were mapped to least one public database. 2 695 unigenes were annotated and assigned to 62 TF families,and 5 920 SSR loci were identified. The majority of mapped unigenes was from Protobothrops mucrosquamatus in the NR database,which revealed their closest homology. Three secretory phospholipase A_2 with different amino acid sequences showed similar spatial structures and all had well-conserved active sites. The 3 D structural models of C-type lectin showed conserved glycosylation binding sites( Asn45). This study will lay the foundation for the further study of the function of snake venom protein,and promoting the development and utilization of genome resources from A. acutus.

[Cloning and characterization of chalcone synthase and chalcone isomerase genes in Arisaema heterophyllum].

Chalcone synthase( CHS) and chalcone isomerase( CHI) are key enzymes in the biosynthesis pathway of flavonoids. In this study,unigenes for CHS and CHI were screened from the transcriptome database of Arisaema heterophyllum. The open reading frame( ORFs) of chalcone synthase( Ah CHS) and chalcone isomerase( Ah CHI) were cloned from the plant by RT-PCR. The physicochemical properties,expression and structure characteristics of the encoded proteins Ah CHS and Ah CHI were analyzed. The ORFs of Ah CHS and Ah CHI were 1 176,630 bp in length and encoded 392,209 amino acids,respectively. Ah CHS functioned as a symmetric homodimer. The N-terminal helix of one monomer entwined with the corresponding helix of another monomer. Each CHS monomer consisted of two structural domains. In particular,four conserved residues define the active site. The tertiary structure of Ah CHI revealed a novel open-faced ß-sandwich fold. A large ß-sheet( ß4-ß11) and a layer of α-helices( α1-α7) comprised the core structure. The residues spanning ß4,ß5,α4,and α6 in the three-dimensional structure were conserved among CHIs from different species. Notably,these structural elements formed the active site on the protein surface,and the topology of the active-site cleft defined the stereochemistry of the cyclization reaction. The homology comparison showed that Ah CHS had the highest similarity to the CHS of Anthurium andraeanum,while Ah CHI had the highest similarity to the CHI of Paeonia delavayi. This study provided the basis for the functional study of Ah CHS and Ah CHI and the further study on plant flavonoid biosynthesis pathway.

Hyperglycemia promotes vasculogenesis in choroidal neovascularization in diabetic mice by stimulating VEGF and SDF-1 expression in retinal pigment epithelial cells.

To investigate the influence of hyperglycemia on the severity of choroidal neovascularization (CNV) in diabetic mice, especially the involvement of bone marrow-derived cells (BMCs) and underlying molecular mechanisms. The mice were randomly divided into control group, diabetes group and diabetes treated with insulin group, which were laser treated to induce CNV. The CNV severity was evaluated by fundus fluorescein angiography, HE staining and choroidal flatmount. The BMCs recruitment and differentiation in CNV were examined in GFP chimeric mice by choroidal flatmount and immunofluorescence. The bone marrow-derived mesenchymal stem cells (BMSCs) recruitment and migration were tested in vivo and in vitro. VEGF and SDF-1 production in vivo and in vitro were tested by realtime PCR and ELISA. The CNV severity and expression of VEGF and SDF-1 were enhanced in DM mice compared with control mice and that insulin treatment decreased CNV severity in DM mice. The DM mice demonstrated more BMCs and bone marrow-derived mesenchymal stem cells (BMSCs) recruited and incorporated into CNV, increased ratio of BMCs expressing endothelial cell marker or macrophage marker, and up-regulated expression of VEGF and SDF-1 in CNV. Human BMSCs migration and expression of VEGF and SDF-1 in retinal pigment epithelial (RPE) cells increased when cultured under high glucose. This study suggested that hyperglycemia enhanced the expression of VEGF and SDF-1 in RPE cells, and promoted recruitment and incorporation of BMCs and affected differentiation of BMCs in CNV, which led to more severe CNV in diabetic mice.

Genomewide analysis indicates that queen larvae have lower methylation levels in the honey bee (Apis mellifera).

The honey bee is a social insect characterized by caste differentiation, by which a young larva can develop into either a queen or a worker. Despite possessing the same genome, queen and workers display marked differences in reproductive capacity, physiology, and behavior. Recent studies have shown that DNA methylation plays important roles in caste differentiation. To further explore the roles of DNA methylation in this process, we analyzed DNA methylome profiles of both queen larvae (QL) and worker larvae (WL) of different ages (2, 4, and 6 day old), by using methylated DNA immunoprecipitation-sequencing (meDIP-seq) technique. The global DNA methylation levels varied between the larvae of two castes. DNA methylation increased from 2-day- to 4-day-old QL and then decreased in 6-day-old larvae. In WL, methylation levels increased with age. The methylcytosines in both larvae were enriched in introns, followed by coding sequence (CDS) regions, CpG islands, 2 kbp downstream and upstream of genes, and 5' and 3' untranslated regions (UTRs). The number of differentially methylated genes (DMGs) in 2-, 4-, and 6-day-old QL and WL was 725, 3,013, and 5,049, respectively. Compared to 4- and 6-day-old WL, a large number of genes in QL were downmethylated, which were involved in many processes including development, reproduction, and metabolic regulation. In addition, some DMGs were concerned with caste differentiation.

Association between genetic variants of the VEGFR-2 gene and the risk of developing endometriosis in Northern Chinese Women.

AIM: To investigate the association of tag single nucleotide polymorphisms (SNPs) in the vascular endothelial growth factor receptor 2 (VEGFR-2) gene with susceptibility to endometriosis. METHODS: This study comprised 571 patients with endometriosis and 580 women in the control group. Five tag SNPs in the VEGFR-2 gene were selected using a Haploview program, and those SNPs were genotyped by a method of polymerase chain reaction and ligase detection reaction. RESULTS: Statistical results show that there was a significant difference in the genotype and allele distribution of the 1192C/T polymorphism between the disease group and the control group (p = 0.041 and 0.017). The women carrying the T allele (C/T+T/T genotype) had a lower risk of developing endometriosis compared with the women with the C/C genotype (OR 0.75, 95% CI 0.57-0.99). There was no significant difference in the allele and genotype distribution of four other tag SNPs (1719T/A, +31C/T, IVS25-92A/G and IVS6+​54C/T) between the disease group and the control group (all p > 0.05). CONCLUSIONS: Our results suggested that the 1192C/T polymorphisms on the VEGFR-2 gene might affect the risk of developing endometriosis in Northern Chinese women of Han ethnicity.

Sebaceous Carcinoma of the Submandibular Gland a Case Report and Review of the Literature.

Objective: Sebaceous carcinoma (SC) of the submandibular gland is extremely rare. Owing to the low morbidity and nonspecific clinical manifestations, diagnosis is commonly delayed, which increases metastasis and mortality. To date, there have been five reported cases of SC of the submandibular gland. Here, we present a new case and review the relevant literature. Methods and Results: A 36-year-old woman presented with an enlarged left submandibular gland. Clinical features included a non-tender solitary nodular mass with normal overlying skin. There were no special findings on computed tomography or ultrasound examination except for a swollen mass in the left submandibular gland. The patient underwent surgical resection. Pathological examination confirmed the diagnosis of SC with nerve infiltration. Immunohistochemical examination of this case showed positive staining for P63, P40, CK7, CK8/18, MLH1, MSH2, MSH6, and PMS2. The specimen was negative for androgen receptor, CEA, S-100, CK5/6, SOX-10, SOX-11, SMA, and GCDFP-15. The KI-67 labeling index was determined to be 15%. PAS and anti-epithelial membrane antigen were positive in partial area. The patient is still undergoing follow-up, and no metastasis or recurrence has been observed for 2 months. Conclusion: This case highlighted the fact that despite its rarity, SC should be considered as a differential diagnosis for masses located in the head and face. Early and accurate diagnosis, followed by wide surgical excision, has a favorable prognosis. Therefore, clinicians should be familiar with the clinical and pathological features of this disease.

Safety and efficacy of a modified busulfan/cyclophosphamide conditioning regimen incorporating cladribine for autologous hematopoietic stem cell transplantation in acute myeloid leukemia.

This is a small phase I study examining the safety and efficacy of a cladribine (CLAD)-containing conditioning regimen prior to autologous hematopoietic stem cell transplantion (auto-HSCT) for patients with acute myeloid leukemia (AML). All patients, aged 15-54 years (median 32 years), had favorable/intermediate risk AML (n = 20) or acute promyelocytic leukemia (APL; n = 2) and no evidence of minimal residual disease (MRD) prior to transplantation. Fourteen of the 22 patients received the conditioning regimen as follows: busulfan (Bu) + cyclophosphamide (Cy) + CLAD + cytarabine (Ara-c) or idarubicin. The conditioning regimen of 8 patients was without Cy nor idarubicin to reducing adverse cardiac reaction: the regimen of Bu + CLAD+ Ara-c for 6 patients; and the regimen of Bu + melphalan + CLAD + Ara-c for the other 2 patients. All 22 AML patients received peripheral blood stem cell transplantation. The number of infused mononuclear cells and CD34+ cells was 10.00 (2.88-20.97) × 108/kg and 1.89 (1.52-10.44) × 106/kg, respectively. Hematopoietic reconstitution was achieved in all patients, with a median time of 13 (10-34) days for neutrophils and 28 (14-113) days for platelets. Two patients suffered from pulmonary infection, 4 patients suffered from septicemia during the neutropenic stage, and the others suffered from infection or gastrointestinal reaction without exceeding grade 3 after conditioning, which were all alleviated by anti-infection and other supportive treatment. None of the patients died of transplantation-related complications. At a median follow-up of 29.5 (ranging from 4.0 to 60.0) months, three patients relapsed after auto-HSCT at a median time of 6 (ranging from 0.5 to 10.0) months. One patient died due to relapse at 18 months after auto-HSCT. The remaining 21 patients were all alive, including 19 patients with negative MRD. The other 2 patients achieved negative MRD after allogeneic HSCT or chemotherapy. The estimated 2-year survival, relapse, and Leukemia-free survival rates were 94.1 ± 5.7%, 14.7 ± 7.9% and 85.3 ± 7.9%, respectively. A CLAD-combination conditioning regimen is efficient and safe for auto-HSCT, indicating an effective approach for AML treatment.

Acute Myeloid Leukemia With CEBPA Mutations: Current Progress and Future Directions.

Mutations in CCAAT enhancer binding protein A gene (CEBPA) are one of the common genetic alterations in acute myeloid leukemia (AML). Recently, the emergence of new evidence makes it necessary to reconsider the subsets and treatment of AML patients with CEBPA mutations. This review will summarize the history of research progress of CEBPA mutations in AML, the heterogeneities of AML with CEBPA double mutations (CEBPA dm), and two special subtypes of CEBPA mutated AML. We will discuss the treatment of AML with CEBPA mutations as well, and finally propose a new algorithm for the treatment of these patients, including both familial and sporadic CEBPA mutated AML patients. This review may be beneficial for further investigation and optimizing clinical management of AML patients with CEBPA mutations.

System pharmacology analysis to decipher the effect and mechanism of active ingredients combination from herb couple on rheumatoid arthritis in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Traditional herb couple Angelicae pubescentis radix (APR) and Notopterygii rhizoma et radix (NRR), composition of two traditional Chinese medicinal herbs, has been used clinically in China for the treatment of rheumatoid arthritis (RA) over years. APR and NRR contain coumarins and phenolic acids, which have been reported to have analgesic and anti-inflammatory activities. AIM OF THE STUDY: The active ingredients combination (AIC) and potential therapeutic mechanism of APR and NRR (AN) herb couple remain unclear. Therefore, the present study aimed to identify the AIC and elucidate the underlying mechanism of AIC on RA. MATERIALS AND METHODS: Firstly, a novel strategy of in vitro experiments, computational analysis, UPLC-QTOF-MS and UPLC-QQQ-MS was established to confirm the optimum ratio of AN herb couple samples and identified the AIC. Then, the anti-arthritis effects of the optimal herb couple and AIC were studied with Collagen II induced rheumatoid arthritis (CIA) rats in vivo. Finally, an integrated model of network pharmacology, metabolomics, gut microbiota analysis and biological techniques were applied to clarify the underlying mechanism through a comprehensive perspective. RESULTS: AN7:3 herb couple was regarded as the optimal ratio of AN herbal samples, and AIC was screened as osthole, columbianadin, notopterol, isoimperatorin, psoralen, xanthotoxin, bergapten, nodakenin and bergaptol respectively. Additionally, AIC exerted similar therapeutic effects as AN 7:3 in CIA rats. Moreover, AIC ameliorated RA might via regulating MAPK signaling pathway, altering metabolic disorders and gut microbiome involved autoimmunity. CONCLUSIONS: our findings provided scientific evidence to support that AIC of AN herb couple could be used as a prebiotic agent for RA. Importantly, this research provided a systematic and feasible strategy to optimize the proportion of medicinal materials and screen AIC from multi-component traditional Chinese herb couples or Chinese medicine formulae. Moreover, it provided a comprehensive perspective to discover AIC, clarify the overall effects and understand the mechanisms for natural products through the perspective of database and multi-omics integration.

Network Pharmacology-Based Analysis on the Effects and Mechanism of the Wang-Bi Capsule for Rheumatoid Arthritis and Osteoarthritis.

Wang-Bi capsule (WB) is a traditional Chinese medicine (TCM)-based herbal formula, and it has been used in the treatment of rheumatoid arthritis (RA) in China for many years. Additionally, WB is also used as a supplement to the treatment of osteoarthritis (OA) in clinical practice. Our research aimed to reveal the therapeutic effects and underling mechanism of WB on RA and OA through computational system pharmacology analysis and experimental study. Based on network pharmacology analysis, a total of 173 bioactive compounds interacted with 417 common gene targets related to WB, RA, and OA, which mainly involved the PI3K-Akt signaling pathway. In addition, the serine-threonine protein kinase 1 (AKT1) might be a core gene protein for the action of WB, which was further emphasized by molecular docking. Moreover, the anti-inflammatory activity of WB in vitro was confirmed by reducing NO production in lipopolysaccharide (LPS)-induced RAW264.7 cells. The anti-RA and OA effects of WB in vivo were confirmed by ameliorating the disease symptoms of collagen II-induced RA (CIA) and monosodium iodoacetate-induced OA (MIA) in rats, respectively. Furthermore, the role of the PI3K-Akt pathway in the action of WB was preliminarily verified by western blot analysis. In conclusion, our study elucidated that WB is a potentially effective strategy for the treatment of RA and OA, which might be achieved by regulating the PI3K-Akt pathway. It provides us with systematic insights into the effects and mechanism of WB on RA and OA.

Tight junction protein 1 promotes vasculature remodeling via regulating USP2/TWIST1 in bladder cancer.

Bladder cancer (BLCA) is the most common malignant tumor of the urinary system and is characterized by high metastatic rates and poor prognosis. The expression of tight junction protein 1 (TJP1) is associated with bladder cancer invasion; however, the mechanism by which TJP1 affects vasculature remodeling remains unknown. In this study, we found that TJP1 expression correlated with tumor angiogenesis and poor overall survival in clinical samples. Furthermore, TJP1 overexpression promoted tumor angiogenesis in BLCA cells and stimulated recruitment of macrophages to tumors by upregulating CCL2 expression. Mechanistically, TJP1 interacted with TWIST1 and enhanced the transcriptional activity of CCL2. The impairment of tumor angiogenesis caused by knockdown of TJP1 was dramatically rescued by overexpression of TWIST1. Furthermore, TJP1 recruited USP2, which deubiquitinated TWIST1, thereby protecting TWIST1 from proteasome-mediated protein degradation. In conclusion, our results suggest that TJP1 controls angiogenesis in BLCA via TWIST1-dependent regulation of CCL2. We demonstrate that TJP1 functions as a scaffold for the interaction between USP2 and TWIST1 and this may provide potential therapeutic targets in bladder cancer.