Aspartoacylase suppresses prostate cancer progression by blocking LYN activation.

BACKGROUND: Globally, despite prostate cancer (PCa) representing second most prevalent malignancy in male, the precise molecular mechanisms implicated in its pathogenesis remain unclear. Consequently, elucidating the key molecular regulators that govern disease progression could substantially contribute to the establishment of novel therapeutic strategies, ultimately advancing the management of PCa. METHODS: A total of 49 PCa tissues and 43 adjacent normal tissues were collected from January 2017 to December 2021 at Zhongnan Hospital of Wuhan University. The advanced transcriptomic methodologies were employed to identify differentially expressed mRNAs in PCa. The expression of aspartoacylase (ASPA) in PCa was thoroughly evaluated using quantitative real-time PCR and Western blotting techniques. To elucidate the inhibitory role of ASPA in PCa cell proliferation and metastasis, a comprehensive set of in vitro and in vivo assays were conducted, including orthotopic and tumor-bearing mouse models (n = 8 for each group). A combination of experimental approaches, such as Western blotting, luciferase assays, immunoprecipitation assays, mass spectrometry, glutathione S-transferase pull-down experiments, and rescue studies, were employed to investigate the underlying molecular mechanisms of ASPA's action in PCa. The Student's t-test was employed to assess the statistical significance between two distinct groups, while one-way analysis of variance was utilized for comparisons involving more than two groups. A two-sided P value of less than 0.05 was deemed to indicate statistical significance. RESULTS: ASPA was identified as a novel inhibitor of PCa progression. The expression of ASPA was found to be significantly down-regulated in PCa tissue samples, and its decreased expression was independently associated with patients' prognosis (HR = 0.60, 95% CI 0.40-0.92, P = 0.018). Our experiments demonstrated that modulation of ASPA activity, either through gain- or loss-of-function, led to the suppression or enhancement of PCa cell proliferation, migration, and invasion, respectively. The inhibitory role of ASPA in PCa was further confirmed using orthotopic and tumor-bearing mouse models. Mechanistically, ASPA was shown to directly interact with the LYN and inhibit the phosphorylation of LYN as well as its downstream targets, JNK1/2 and C-Jun, in both PCa cells and mouse models, in an enzyme-independent manner. Importantly, the inhibition of LYN activation by bafetinib abrogated the promoting effect of ASPA knockdown on PCa progression in both in vitro and in vivo models. Moreover, we observed an inverse relationship between ASPA expression and LYN activity in clinical PCa samples, suggesting a potential regulatory role of ASPA in modulating LYN signaling. CONCLUSION: Our findings provide novel insights into the tumor-suppressive function of ASPA in PCa and highlight its potential as a prognostic biomarker and therapeutic target for the management of this malignancy.

Using Haemocoagulase Agkistrodon in Patients Undergoing Transurethral Plasmakinetic Resection of the Prostate: A Pilot, Real-World, and Propensity Score-Matched Study.

Objectives: To compare the clinical outcomes of using different hemostatic agents after transurethral plasmakinetic resection of the prostate (TUPKP) in benign prostatic hyperplasia (BPH) patients. Methods: The patients were divided into 5 groups according to the hemostatic agents used after TUPKP, including the haemocoagulase agkistrodon for injection (HCA), hemocoagulase for injection (HC), hemocoagulase bothrops atrox for injection (HCB), ethylenediamine diaceturate injection (EDD), and tranexamic acid (TXA). Propensity score matching was performed based on age, body mass index, prostate volume, hypertension status, fasting blood glucose, smoking, and drinking history. The hospitalization time, bladder irrigation time, indwelling catheterization time, the patency of urine flow, and blood transfusion records were used as outcome indicators to compare the clinical effects of these five agents. Results: We finally matched 65 pairs receiving HCA or HC, 71 pairs receiving HCA or HCB, 38 pairs receiving HCA or TXA, and 29 pairs receiving HCA or EDD. Compared with HC, HCA given during the perioperative period significantly reduced the median hospitalization time [7.00 days (5.00, 8.00) vs. 9.00 days (8.00, 10.00); p < 0.001] and median catheterization time (109.00 hours [88.00, 129.00] vs. 164.00 hours [114.00, 189.00], p < 0.001). Compared with EDD, the median hospitalization time (7.00 days [6.00, 8.00] vs. 10.00 days [8.00, 11.00]; p < 0.001) and median catheterization time (113.00 hours [95.00, 143.00] vs. 160.00 hours [139.00, 168.00]; p < 0.001) were also significant shorter in HCA group. Compared with HCB, median bladder irrigation time (45.00 hours [27.00, 71.00] vs. 49.00 hours [45.00, 72.00]; p = 0.04) was shorter in the HCA group. However, there were no statistical differences in outcomes between HCA and TXA. Conclusions: HCA probably has an advantage over HC, HCB, and EDD in reducing the hospitalization time, catheterization time, and bladder irrigation time among BPH patients undergoing TUPKP.

Cloning of a SEPALLATA4-like gene (IiSEP4) in Isatis indigotica Fortune and characterization of its function in Arabidopsis thaliana.

E-class MADS-box genes, SEPALLATA (SEP), participate in various aspects of plant development together with B-, C- and D-class MADS-box genes. IiSEP4, a homologous gene of SEP4, was cloned from Isatis indigotica. IiSEP4 was highly expressed in sepals, and its mRNA was mildly detected in leaves, inflorescences, flowers, stamens and young silicles. Constitutive expression of IiSEP4 in Arabidopsis thaliana caused early flowering, accompanied by the reduction of flowers and floral organs. Moreover, the sepals in some flowers were transformed into carpelloid structures with stigmatic papillae, and obviously accompanied by ovule formation. Yeast two-hybrid assays demonstrated that IiSEP4 interacts with other woad MADS proteins to determine the identity of floral organs. These findings reveal the important roles of IiSEP4 in floral development of I. indigotica. The results of this study can lay a foundation for further study on biological functions of MADS transcriptional factors in I. indigotica.

Ectopic expression of IiSHP2 from Isatis indigotica Fortune, a PLE-lineage MADS-box gene, influences leaf, floral organ and silique morphology in Arabidopsis thaliana.

In order to ascertain the regulatory mechanism of fruit development in Isatis indigotica Fortune, the complementary DNA (cDNA) sequence of the SHATTERPROOF 2 (SHP2) orthologous gene was identified by Rapid Amplification of cDNA Ends technology and the corresponding gene was named IiSHP2. The expression pattern of IiSHP2 was determined by quantitative reverse transcription-polymerase chain reaction and wild-type Col-0 Arabidopsis plants were transformed with the IiSHP2 gene using Agrobacterium tumefaciens and the floral-dip method. Expression analyses indicated that IiSHP2 was highly expressed in flowers, silicles and seeds. Compared to wild-type plants, IiSHP2 transgenic lines bolted earlier. Detailed phenotypic observations showed that the size of the rosette and cauline leaves in transgenic lines was reduced and the cauline leaves of the transgenic lines were incurved and displayed a funnel-like shape. During the reproductive growth stage, IiSHP2 transgenic plants produced shortened sepals and the flower buds were not encapsulated completely. Moreover, the petals of the transgenic lines were converted into stamineous tissues, accompanied by exposed stamens, short malformed siliques and wrinkled valves, indicating a severe decline in fertility. These experimental conclusions are valuable as a reference for the breeding of medicinal plants.

Resveratrol induces apoptosis of benign prostatic hyperplasia epithelial cell line (BPH-1) through p38 MAPK-FOXO3a pathway.

BACKGROUND: Resveratrol is reported to inhibit the growth of prostate, which is characteristic of benign prostatic hyperplasia (BPH) condition. However, the mechanism remains unclear. This study aimed to identify the effects and probable mechanism of resveratrol on BPH. METHODS: We used the BPH epithelial cell line BPH-1 to investigate the effect of resveratrol. Cells were treated with various concentrations of resveratrol, and its effects on cells viability, apoptosis, ROS accumulation, and cell cycle were assessed. Western blot was used to examine activation of p38 MAPK and protein levels of FOXO3a, Bcl2, Bcl-XL, and caspase3. Cells were also co-treated with the p38 MAPK inhibitor SB203580 or ROS scavenger N-Acetyl-L-cysteine (NAC) to further investigate the mechanism. RESULTS: Resveratrol treatment inhibited the growth of BPH-1 and increased apoptosis of cells. In addition, levels of phosphorylated p38 MAPK level was elevated and FOXO3a repression was observed. Concomitantly, ROS was accumulated. All of these resveratrol-mediated effects were suppressed by additional treatment with SB203580 or NAC. Resveratrol was also found to induce cell cycle arrest at S phase. CONCLUSIONS: Resveratrol can activate p38 MAPK and repress FOXO3a, thereby causing repression of SOD2, catalase, and increase of ROS accumulation, leading to apoptosis in BPH-1 cells.

A SEPALLATA1-like gene of Isatis indigotica Fort. regulates flowering time and specifies floral organs.

An orthologous gene of SEPALLATA1, designated as IiSEP1, was isolated from Isatis indigotica. The genomic DNA of IiSEP1 is 3.1 Kb in length. The full-length cDNA of IiSEP1 is 1481 bp and contains a 756 bp ORF encoding a 251-amino-acid protein. Sequence comparison revealed that IiSEP1 belonged to the MADS-box gene family. IiSEP1 contains 7 exons and 6 introns, showing similar exon-intron structure with Arabidopsis SEP1. Phylogenetic analysis suggested that IiSEP1 belonged to AGL2/SEP subfamily and was likely to be an I. indigotica ortholog of Arabidopsis SEP1. Quantitative real-time PCR showed that IiSEP1 was predominantly expressed in the reproductive organs. Ectopic expression of IiSEP1 in Arabidopsis resulted in early flowering, accompanied with the reduction of inflorescence number and the production of terminal flower on the top of the main stems. Moreover, IiSEP1 overexpressing flowers generated numerous variations in phenotype. The sepals were changed into petal-sepal mosaic structures or displayed carpelloid features, and transparent ovules were formed in internal surface of these sepals. In addition, some flowers were constituted by sepals and pistil, but lacked petals and stamens. Taken together, IiSEP1 might play important roles in reproductive growth of I. indigotica and could affect the morphogenesis of flowers and fruits.

Constitutive expression of NtabSPL6-1 in tobacco and Arabidopsis could change the structure of leaves and promote the development of trichomes.

The coding sequence of NtabSPL6-1 was cloned by high-fidelity PCR with specific primers and was used in construction of a binary vector for overexpression. Wild-type Col-0 Arabidopsis plants and Qinyan95 tobacco leaves were transformed using floral dip and leaf disc methods, respectively. Phenotypic observation showed that constitutive expression of NtabSPL6-1 in Arabidopsis could promote the development of trichomes on leaf epidermis and influence the growth pattern of cauline leaves. In tobacco, ectopic expression of NtabSPL6-1 led to dwarfism of the plants and alteration of the leaf structure, accompanied by changes of the glandular trichomes in development. At the same time, the self-regulation capability of NtabSPL6-1 was determined by yeast two-hybrid system. The results indicated that SBP-C terminal domain and C terminal domain of NtabSPL6-1 possessed strong transcriptional activation ability; the intact protein, N terminal domain, and the first peptide fragment in N terminal domain possessed weak transcriptional activation ability; and the second and the third peptide fragments in N terminal domain had no transcriptional activation ability, suggesting the N terminal domain of NtabSPL6-1 could block the activity of the C terminal domain. NtabSPL6-1 may affect the resistance of plants to biotic stress factors indirectly by regulation of the trichome growth.