Structure-based identification of a G protein-biased allosteric modulator of cannabinoid receptor CB1.

Cannabis sativa is known for its therapeutic benefit in various diseases including pain relief by targeting cannabinoid receptors. The primary component of cannabis, Δ9-tetrahydrocannabinol (THC), and other agonists engage the orthosteric site of CB1, activating both Gi and ß-arrestin signaling pathways. The activation of diverse pathways could result in on-target side effects and cannabis addiction, which may hinder therapeutic potential. A significant challenge in pharmacology is the design of a ligand that can modulate specific signaling of CB1. By leveraging insights from the structure-function selectivity relationship (SFSR), we have identified Gi signaling-biased agonist-allosteric modulators (ago-BAMs). Further, two cryoelectron microscopy (cryo-EM) structures reveal the binding mode of ago-BAM at the extrahelical allosteric site of CB1. Combining mutagenesis and pharmacological studies, we elucidated the detailed mechanism of ago-BAM-mediated biased signaling. Notably, ago-BAM CB-05 demonstrated analgesic efficacy with fewer side effects, minimal drug toxicity and no cannabis addiction in mouse pain models. In summary, our finding not only suggests that ago-BAMs of CB1 provide a potential nonopioid strategy for pain management but also sheds light on BAM identification for GPCRs.

Identifying Patients With Rapid Progression From Hormone-Sensitive to Castration-Resistant Prostate Cancer: A Retrospective Study.

Prostate cancer (PCa) is the second most prevalent malignancy and the fifth cause of cancer-related deaths in men. A crucial challenge is identifying the population at risk of rapid progression from hormone-sensitive prostate cancer (HSPC) to lethal castration-resistant prostate cancer (CRPC). We collected 78 HSPC biopsies and measured their proteomes using pressure cycling technology and a pulsed data-independent acquisition pipeline. We quantified 7355 proteins using these HSPC biopsies. A total of 251 proteins showed differential expression between patients with a long- or short-term progression to CRPC. Using a random forest model, we identified seven proteins that significantly discriminated long- from short-term progression patients, which were used to classify PCa patients with an area under the curve of 0.873. Next, one clinical feature (Gleason sum) and two proteins (BGN and MAPK11) were found to be significantly associated with rapid disease progression. A nomogram model using these three features was generated for stratifying patients into groups with significant progression differences (p-value = 1.3×10-4). To conclude, we identified proteins associated with a fast progression to CRPC and an unfavorable prognosis. Based on these proteins, our machine learning and nomogram models stratified HSPC into high- and low-risk groups and predicted their prognoses. These models may aid clinicians in predicting the progression of patients, guiding individualized clinical management and decisions.

The C. elegans TspanC8 tetraspanin TSP-14 exhibits isoform-specific localization and function.

Tetraspanin proteins are a unique family of highly conserved four-pass transmembrane proteins in metazoans. While much is known about their biochemical properties, the in vivo functions and distribution patterns of different tetraspanin proteins are less understood. Previous studies have shown that two paralogous tetraspanins that belong to the TspanC8 subfamily, TSP-12 and TSP-14, function redundantly to promote both Notch signaling and bone morphogenetic protein (BMP) signaling in C. elegans. TSP-14 has two isoforms, TSP-14A and TSP-14B, where TSP-14B has an additional 24 amino acids at its N-terminus compared to TSP-14A. By generating isoform specific knock-ins and knock-outs using CRISPR, we found that TSP-14A and TSP-14B share distinct as well as overlapping expression patterns and functions. While TSP-14A functions redundantly with TSP-12 to regulate body size and embryonic and vulva development, TSP-14B primarily functions redundantly with TSP-12 to regulate postembryonic mesoderm development. Importantly, TSP-14A and TSP-14B exhibit distinct subcellular localization patterns. TSP-14A is localized apically and on early and late endosomes. TSP-14B is localized to the basolateral cell membrane. We further identified a di-leucine motif within the N-terminal 24 amino acids of TSP-14B that serves as a basolateral membrane targeting sequence, and showed that the basolateral membrane localization of TSP-14B is important for its function. Our work highlights the diverse and intricate functions of TspanC8 tetraspanins in C. elegans, and demonstrates the importance of dissecting the functions of these important proteins in an intact living organism.

KLHL21 suppresses gastric tumourigenesis via maintaining STAT3 signalling equilibrium in stomach homoeostasis.

OBJECTIVE: Precancerous metaplasia transition to dysplasia poses a risk for subsequent intestinal-type gastric adenocarcinoma. However, the molecular basis underlying the transformation from metaplastic to cancerous cells remains poorly understood. DESIGN: An integrated analysis of genes associated with metaplasia, dysplasia was conducted, verified and characterised in the gastric tissues of patients by single-cell RNA sequencing and immunostaining. Multiple mouse models, including homozygous conditional knockout Klhl21-floxed mice, were generated to investigate the role of Klhl21 deletion in stemness, DNA damage and tumour formation. Mass-spectrometry-based proteomics and ribosome sequencing were used to elucidate the underlying molecular mechanisms. RESULTS: Kelch-like protein 21 (KLHL21) expression progressively decreased in metaplasia, dysplasia and cancer. Genetic deletion of Klhl21 enhances the rapid proliferation of Mist1+ cells and their descendant cells. Klhl21 loss during metaplasia facilitates the recruitment of damaged cells into the cell cycle via STAT3 signalling. Increased STAT3 activity was confirmed in cancer cells lacking KLHL21, boosting self-renewal and tumourigenicity. Mechanistically, the loss of KLHL21 promotes PIK3CB mRNA translation by stabilising the PABPC1-eIF4G complex, subsequently causing STAT3 activation. Pharmacological STAT3 inhibition by TTI-101 elicited anticancer effects, effectively impeding the transition from metaplasia to dysplasia. In patients with gastric cancer, low levels of KLHL21 had a shorter survival rate and a worse response to adjuvant chemotherapy. CONCLUSIONS: Our findings highlighted that KLHL21 loss triggers STAT3 reactivation through PABPC1-mediated PIK3CB translational activation, and targeting STAT3 can reverse preneoplastic metaplasia in KLHL21-deficient stomachs.

Unnatural Peptide Assemblies Rapidly Deplete Cholesterol and Potently Inhibit Cancer Cells.

Cholesterol-rich membranes play a pivotal role in cancer initiation and progression, necessitating innovative approaches to target these membranes for cancer inhibition. Here we report the first case of unnatural peptide (1) assemblies capable of depleting cholesterol and inhibiting cancer cells. Peptide 1 self-assembles into micelles and is rapidly taken up by cancer cells, especially when combined with an acute cholesterol-depleting agent (MßCD). Click chemistry has confirmed that 1 depletes cell membrane cholesterol. It localizes in membrane-rich organelles, including the endoplasmic reticulum, Golgi apparatus, and lysosomes. Furthermore, 1 potently inhibits malignant cancer cells, working synergistically with cholesterol-lowering agents. Control experiments have confirmed that C-terminal capping and unnatural amino acid residues (i.e., BiP) are essential for both cholesterol depletion and potent cancer cell inhibition. This work highlights unnatural peptide assemblies as a promising platform for targeting the cell membrane in controlling cell fates.

NOTCH3 promotes docetaxel resistance of prostate cancer cells through regulating TUBB3 and MAPK signaling pathway.

Docetaxel is the preferred chemotherapeutic agent in patients with castrate-resistant prostate cancer (CRPC). However, patients eventually develop docetaxel resistance and in the absence of effective treatment options. Consequently, it is essential to investigate the mechanisms generating docetaxel resistance and develop novel alternative therapeutic targets. RNA sequencing was undertaken on docetaxel-sensitive and docetaxel-resistant prostate cancer (PCa) cells. Subsequently, chemoresistance, cancer stemness, and lipid metabolism were investigated. To obtain insight into the precise activities and action mechanisms of NOTCH3 in docetaxel-resistant PCa, immunoprecipitation, mass spectrometry, ChIP, luciferase reporter assay, cell metabolism, and animal experiments were performed. Through RNA sequencing analysis, we found that NOTCH3 expression was markedly higher in docetaxel-resistant cells relative to parental cells, and that this trend was continued in docetaxel-resistant PCa tissues. Experiments in vitro and in vivo revealed that NOTCH3 enhanced stemness, lipid metabolism, and docetaxel resistance in PCa. Mechanistically, NOTCH3 is bound to TUBB3 and activates the MAPK signaling pathway. Moreover, NOTCH3 was directly regulated by MEF2A in docetaxel-resistant cells. Notably, targeting NOTCH3 and the MEF2A/TUBB3 signaling axis was related to docetaxel chemoresistance in PCa. Overall, these results demonstrated that NOTCH3 fostered stemness, lipid metabolism, and docetaxel resistance in PCa via the TUBB3 and MAPK signaling pathways. Therefore, NOTCH3 may be employed as a prognostic biomarker in PCa patients. NOTCH3 could be a therapeutic target for PCa patients, particularly those who have developed docetaxel resistance.

SOX9 Modulates the Transformation of Gastric Stem Cells Through Biased Symmetric Cell Division.

BACKGROUND & AIMS: Transformation of stem/progenitor cells has been associated with tumorigenesis in multiple tissues, but stem cells in the stomach have been hard to localize. We therefore aimed to use a combination of several markers to better target oncogenes to gastric stem cells and understand their behavior in the initial stages of gastric tumorigenesis. METHODS: Mouse models of gastric metaplasia and cancer by targeting stem/progenitor cells were generated and analyzed with techniques including reanalysis of single-cell RNA sequencing and immunostaining. Gastric cancer cell organoids were genetically manipulated with clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) for functional studies. Cell division was determined by bromodeoxyuridine-chasing assay and the assessment of the orientation of the mitotic spindles. Gastric tissues from patients were examined by histopathology and immunostaining. RESULTS: Oncogenic insults lead to expansion of SOX9+ progenitor cells in the mouse stomach. Genetic lineage tracing and organoid culture studies show that SOX9+ gastric epithelial cells overlap with SOX2+ progenitors and include stem cells that can self-renew and differentiate to generate all gastric epithelial cells. Moreover, oncogenic targeting of SOX9+SOX2+ cells leads to invasive gastric cancer in our novel mouse model (Sox2-CreERT;Sox9-loxp(66)-rtTA-T2A-Flpo-IRES-loxp(71);Kras(Frt-STOP-Frt-G12D);P53R172H), which combines Cre-loxp and Flippase-Frt genetic recombination systems. Sox9 deletion impedes the expansion of gastric progenitor cells and blocks neoplasia after Kras activation. Although Sox9 is not required for maintaining tissue homeostasis where asymmetric division predominates, loss of Sox9 in the setting of Kras activation leads to reduced symmetric cell division and effectively attenuates the Kras-dependent expansion of stem/progenitor cells. Similarly, Sox9 deletion in gastric cancer organoids reduces symmetric cell division, organoid number, and organoid size. In patients with gastric cancer, high levels of SOX9 are associated with recurrence and poor prognosis. CONCLUSION: SOX9 marks gastric stem cells and modulates biased symmetric cell division, which appears to be required for the malignant transformation of gastric stem cells.

Freeze Metal Halide Perovskite for Dramatic Laser Tuning: Direct Observation via In Situ Cryo-Electron Microscope.

A frozen-temperature (below -28 °C) laser tuning way is developed to optimize metal halide perovskite (MHP)'s stability and opto-electronic properties, for emitter, photovoltaic and detector applications. Here freezing can adjust the competitive laser irradiation effects between damaging and annealing/repairing. And the ligand shells on MHP surface, which are widely present for many MHP materials, can be frozen and act as transparent solid templates for MHP's re-crystallization/re-growth during the laser tuning. With model samples of different types of CsPbBr3 nanocube arrays,an attempt is made to turn the dominant exposure facet from low-energy [100] facet to high-energy [111], [-211], [113] and [210] ones respectively; selectively removing the surface impurities and defects of CsPbBr3 nanocubes to enhance the irradiation durability by 101 times; and quickly (tens of seconds) modifying a Ruddlesden-Popper (RP) boundary into another type of boundary like twinning, and so on. The laser tuning mechanism is revealed by an innovative in situ cryo-transmission electron microscope (cryo-TEM) exploration at atomic resolution.

Enzyme-Instructed Intracellular Peptide Assemblies.

Higher-order or supramolecular protein assemblies, usually regulated by enzymatic reactions, are ubiquitous and essential for cellular functions. This evolutionary fact has provided a rigorous scientific foundation, as well as an inspiring blueprint, for exploring supramolecular assemblies of man-made molecules that are responsive to biological cues as a novel class of therapeutics for biomedicine. Among the emerging man-made supramolecular structures, peptide assemblies, formed by enzyme reactions or other stimuli, have received most of the research attention and advanced most rapidly.In this Account, we will review works that apply enzyme-instructed self-assembly (EISA) to generate intracellular peptide assemblies for developing a new kind of biomedicine, especially in the field of novel cancer nanomedicines and modulating cell morphogenesis. As a versatile and cell-compatible approach, EISA can generate nondiffusive peptide assemblies locally; thus, it provides a unique approach to target subcellular organelles with exceptional cell selectivity. We have arranged this Account in the following way: after introducing the concept, simplicity, and uniqueness of EISA, we discuss the EISA-formed intracellular peptide assemblies, including artificial filaments, in the cell cytosol. Then, we describe the representative examples targeting subcellular organelles, such as mitochondria, endoplasmic reticulum, Golgi apparatus, lysosomes, and the nucleus, by enzyme-instructed intracellular peptide assemblies for potential cancer therapeutics. After that, we highlight the recent exploration of the transcytosis of peptide assemblies for controlling cell morphogenesis. Finally, we provide a brief outlook of enzyme-instructed intracellular peptide assemblies. This Account aims to illustrate the promise of EISA-generated intracellular peptide assemblies in understanding diseases, controlling cell behaviors, and developing new therapeutics from a class of less explored molecular entities, which are substrates of enzymes and become building blocks of self-assembly after the enzymatic reactions.

Accelerating Cellular Uptake with Unnatural Amino Acid for Inhibiting Immunosuppressive Cancer Cells.

Targeting immunosuppressive metastatic cancer cells is a key challenge in therapy. We recently have shown that a rigid-rod aromatic, pBP-NBD, that responds to enzymes and kill immunosuppressive metastatic osteosarcoma (mOS) and castration resistant prostate cancer (CRPC) cells in mimetic bone microenvironment. However, pBP-NBD demonstrated moderate efficacy against CRPC cells. To enhance activity, we incorporated the unnatural amino acid L- or D-4,4'-biphenylalanine (L- or D-BiP) into pBP-NBD, drastically increasing cellular uptake and CRPC inhibition. Specifically, we inserted BiP into pBP-NBD to target mOS (Saos2 and SJSA1) and CRPC (VCaP and PC3) cells with overexpressed phosphatases. Our results show that the D-peptide backbone with an aspartate methyl diester at the C-terminal offers the highest activity against these immunosuppressive mOS and CRPC cells. Importantly, imaging shows that the peptide assemblies almost instantly enter the cells and accumulate primarily within the endoplasmic reticulum of Saos2, SJSA1, and PC3 cells and at the lysosomes of VCaP cells. By using BiP to boost cellular uptake and self-assembly within cancer cells, this work illustrates an unnatural hydrophobic amino acid as a versatile and effective residue to boost endocytosis of synthetic peptides for intracellular self-assembly.

Assessment of the Enzymatic Dephosphorylation Kinetics in the Assemblies of a Phosphopentapeptide that Forms Intranuclear Nanoribbons.

Although the formation of peptide assemblies catalyzed by alkaline phosphatase (ALP) has received increasing attention in inhibiting cancer cells, the detailed enzyme kinetics of the dephosphorylation of the corresponding phosphopeptide assemblies have yet to be determined. We recently discovered that assemblies from a phosphopentapeptide can form intracellular nanoribbons that kill induced pluripotent stem cells or osteosarcoma cells, but the kinetics of enzymatic dephosphorylation remain unknown. Thus, we chose to examine the enzyme kinetics of the dephosphorylation of the phosphopentapeptide [NBD-LLLLpY (1)] from concentrations below to above its critical micelle concentration (CMC). Our results show that the phosphopeptide exhibits a CMC of 75 µM in phosphate saline buffer, and the apparent Vmax and Km values of alkaline phosphatase catalyzed dephosphorylation are approximately 0.24 µM/s and 5.67 mM, respectively. Despite dephosphorylation remaining incomplete at 60 min in all the concentrations tested, dephosphorylation of the phosphopeptide at concentrations of 200 µM or above mainly results in nanoribbons, dephosphorylation at concentrations of CMC largely produces nanofibers, and dephosphorylation below the CMC largely generates nanoparticles. Moreover, the formation of nanoribbons correlates with the intranuclear accumulation of the pentapeptide. By providing the first examination of the enzymatic kinetics of phosphopeptide assemblies, this work further supports the notion that the assemblies of phosphopentapeptides can act as a new functional entity for controlling cell fates.

Impact of chemotherapy delay on long-term prognosis of laparoscopic radical surgery for locally advanced gastric cancer: a pooled analysis of four randomized controlled trials.

BACKGROUND: Adjuvant chemotherapy following curative surgery for locally advanced gastric cancer (AGC) significantly improves long-term patient prognosis. However, delayed chemotherapy (DC), in which patients are unable to receive timely treatment, is a common phenomenon in clinical practice for various reasons. This study aimed to investigate the impact of DC on the prognosis of patients with stage II-III locally AGC and explore the associated risk factors. METHODS: Data from four prospective studies were included in the pooled analysis. The planned chemotherapy (PC) group was defined as the time interval between surgery and the first chemotherapy ≤ 49 d, while the DC group was defined as the time interval between surgery and chemotherapy > 49 d. The prognosis, recurrence, and risk factors were compared, and a nomogram for predicting DC was established. RESULTS: In total, 596 patients were included, of whom 531 (89.1%) had PC and 65 (10.9%) had DC. Survival analysis revealed that the 5-year overall survival (OS) and disease-free survival (DFS) were significantly lower in the DC group than those in the PC group (log-rank P < 0.001). Cox univariable and multivariable analyses showed that DC was an independent risk factor for OS and DFS in stage II-III patients (P < 0.05). Based on the significant factors for DC, a prediction model was established that had a good fit, high accuracy (AUC = 0.780), and clinical applicability in both the training and validation sets. CONCLUSION: Delayed chemotherapy after gastrectomy is associated with poor long-term prognosis in patients with locally advanced stage II-III GC disease. But standardized, full-cycle adjuvant chemotherapy after surgery may play a remedial role, and can to a certain extent compensate the poor effects caused by delayed chemotherapy.

Long-term oncological outcomes of 3D versus 2D laparoscopic gastrectomy for gastric cancer: a randomized clinical trial.

BACKGROUND: Laparoscopy-assisted gastrectomy (LG) is rapidly gaining popularity owing to its minimal invasiveness. Previous studies have found that compared with two-dimensional (2D)-LG, three-dimensional (3D)-LG showed better short-term outcomes. However, the long-term oncological outcomes in patients with locally resectable gastric cancer (GC) remain controversial. METHODS: In this noninferiority, open-label, randomized clinical trial, a total of 438 eligible GC participants were randomly assigned in a 1:1 ratio to either 3D-LG or 2D-LG from January 2015 to April 2016. The primary endpoint was operating time, while the secondary endpoints included 5-year overall survival (OS), disease-free survival (DFS), and recurrence pattern. RESULTS: Data from 401 participants were included in the per-protocol analysis, with 204 patients in the 3D group and 197 patients in the 2D group. The 5-year OS and DFS rates were comparable between the 3D and 2D groups (5-year OS: 70.6% vs. 71.1%, Log-rank P = 0.743; 5-year DFS: 68.1% vs. 69.0%, log-rank P = 0.712). No significant differences were observed between the 3D and 2D groups in the 5-year recurrence rate (28.9% vs. 28.9%, P = 0.958) or recurrence time (mean time, 22.6 vs. 20.5 months, P = 0.412). Further stratified analysis based on the type of gastrectomy, postoperative pathological staging, and preoperative BMI showed that the 5-year OS, DFS, and recurrence rates of the 3D group in each subgroup were similar to those of the 2D group (all P > 0.05). CONCLUSIONS: For patients with locally resectable GC, 3D-LG performed by experienced surgeons in high-volume professional institutions can achieve long-term oncological outcomes comparable to those of 2D-LG. REGISTRATION NUMBER: NCT02327481 ( http://clinicaltrials.gov ).

Oncoprotein LAMTOR5-mediated CHOP silence via DNA hypermethylation and miR-182/miR-769 in promotion of liver cancer growth.

C/EBP homologous protein (CHOP) triggers the death of multiple cancers via endoplasmic reticulum (ER) stress. However, the function and regulatory mechanism of CHOP in liver cancer remain elusive. We have reported that late endosomal/lysosomal adapter, mitogen-activated protein kinase and mTOR activator 5 (LAMTOR5) suppresses apoptosis in various cancers. Here, we show that the transcriptional and posttranscriptional inactivation of CHOP mediated by LAMTOR5 accelerates liver cancer growth. Clinical bioinformatic analysis revealed that the expression of CHOP was low in liver cancer tissues and that its increased expression predicted a good prognosis. Elevated CHOP contributed to destruction of LAMTOR5-induced apoptotic suppression and proliferation. Mechanistically, LAMTOR5-recruited DNA methyltransferase 1 (DNMT1) to the CpG3 region (-559/-429) of the CHOP promoter and potentiated its hypermethylation to block its interaction with general transcription factor IIi (TFII-I), resulting in its inactivation. Moreover, LAMTOR5-enhanced miR-182/miR-769 reduced CHOP expression by targeting its 3'UTR. Notably, lenvatinib, a first-line targeted therapy for liver cancer, could target the LAMTOR5/CHOP axis to prevent liver cancer progression. Accordingly, LAMTOR5-mediated silencing of CHOP via the regulation of ER stress-related apoptosis promotes liver cancer growth, providing a theoretical basis for the use of lenvatinib for the treatment of liver cancer.

Appraisal of surgical outcomes and oncological efficiency of intraoperative adverse events in robotic radical gastrectomy for gastric cancer.

BACKGROUND: Surgical quality control is a crucial determinant of evaluating the tumor efficacy. OBJECTIVE: To assess the ClassIntra grade for quality control and oncological outcomes of robotic radical surgery for gastric cancer (GC). METHODS: Data of patients undergoing robotic radical surgery for GC at a high-volume center were retrospectively analyzed. Patients were categorized into two groups, the intraoperative adverse event (iAE) group and the non-iAE group, based on the occurrence of intraoperative adverse events. The iAEs were further classified into five sublevels (ranging from I to V according to severity) based on the ClassIntra grade. Surgical performance was assessed using the Objective Structured Assessment of Technical Skill (OSATS) and the General Error Reporting Tool. RESULTS: This study included 366 patients (iAE group: n = 72 [19.7%] and non-iAE group: n = 294 [80.3%]). The proportion of ClassIntra grade II patients was the highest in the iAE group (54.2%). In total and distal gastrectomies, iAEs occurred most frequently in the suprapancreatic area (50.0% and 54.8%, respectively). In total gastrectomy, grade IV iAEs were most common during lymph node dissection in the splenic hilum area (once for bleeding [grade IV] and once for injury [grade IV]). The overall survival (OS) and disease-free survival of the non-iAE group were significantly better than those of the iAE group (Log rank P < 0.001). Uni- and multi-variate analyses showed that iAEs were key prognostic indicators, independent of tumor stage and adjuvant chemotherapy (P < 0.001). CONCLUSION: iAEs in patients who underwent robotic radical gastrectomy significantly correlated with the occurrence of postoperative complications and a poor long-term prognosis. Therefore, utilization and inclusion of ClassIntra grading as a crucial surgical quality control and prognostic indicator in the routine surgical quality evaluation system are recommended.

Robot-assisted versus laparoscopic-assisted gastrectomy among malnourished patients with gastric cancer based on textbook outcome.

BACKGROUND: Textbook outcome (TO) has been widely employed as a comprehensive indicator to assess the short-term prognosis of patients with cancer. Preoperative malnutrition is a potential risk factor for adverse surgical outcomes in patients with gastric cancer (GC). This study aimed to compare the TO between robotic-assisted gastrectomy (RAG) and laparoscopic-assisted gastrectomy (LAG) in malnourished patients with GC. METHODS: According to the diagnostic consensus of malnutrition proposed by Global Leadership Initiative on Malnutrition (GLIM) and Nutrition Risk Index (NRI), 895 malnourished patients with GC who underwent RAG (n = 115) or LAG (n = 780) at a tertiary referral hospital between January 2016 and May 2021 were included in the propensity score matching (PSM, 1:2) analysis. RESULTS: After PSM, no significant differences in clinicopathological characteristics were observed between the RAG (n = 97) and LAG (n = 194) groups. The RAG group had significantly higher operative time and lymph nodes harvested, as well as significantly lower blood loss and hospital stay time compared to the LAG group. More patients in the RAG achieved TO. Logistic regression analysis revealed that RAG was an independent protective factor for achieving TO. There were more adjuvant chemotherapy (AC) cycles in the RAG group than in the LAG group. After one year of surgery, a higher percentage of patients (36.7% vs. 22.8%; P < 0.05) in the RAG group recovered from malnutrition compared to the LAG group. CONCLUSIONS: For malnourished patients with GC, RAG performed by experienced surgeons can achieved a higher rate of TO than those of LAG, which directly contributed to better AC compliance and a faster restoration of nutritional status.

Reversed-Phase Medium-Pressure Liquid Chromatography Purification of Omega-3 Fatty Acid Ethyl Esters Using AQ-C18.

Omega-3 fatty acids are in high demand due to their efficacy in treating hypertriglyceridemia and preventing cardiovascular diseases. However, the growth of the industry is hampered by low purity and insufficient productivity. This study aims to develop an efficient RP-MPLC purification method for omega-3 fatty acid ethyl esters with high purity and capacity. The results indicate that the AQ-C18 featuring polar end-capped silanol groups outperformed C18 and others in retention time and impurity separation. By injecting pure fish oil esters with a volume equivalent to a 1.25% bed volume on an AQ-C18 MPLC column using a binary isocratic methanol-water (90:10, v:v) mobile phase at 30 mL/min, optimal omega-3 fatty acid ethyl esters were obtained, with the notable purity of 90.34% and a recovery rate of 74.30%. The total content of EPA and DHA produced increased from 67.91% to 85.27%, meeting the acceptance criteria of no less than 84% set by the 2020 edition of the Pharmacopoeia of the People's Republic of China. In contrast, RP-MPLC significantly enhanced the production efficiency per unit output compared to RP-HPLC. This study demonstrates a pioneering approach to producing omega-3 fatty acid ethyl esters with high purity and of greater quantity using AQ-C18 RP-MPLC, showing this method's significant potential for use in industrial-scale manufacturing.

Targeted Affinity Purification and Mechanism of Action of Angiotensin-Converting Enzyme (ACE) Inhibitory Peptides from Sea Cucumber Gonads.

Protein hydrolysates from sea cucumber (Apostichopus japonicus) gonads are rich in active materials with remarkable angiotensin-converting enzyme (ACE) inhibitory activity. Alcalase was used to hydrolyze sea cucumber gonads, and the hydrolysate was separated by the ultrafiltration membrane to produce a low-molecular-weight peptide component (less than 3 kDa) with good ACE inhibitory activity. The peptide component (less than 3 kDa) was isolated and purified using a combination method of ACE gel affinity chromatography and reverse high-performance liquid chromatography. The purified fractions were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS), and the resulting products were filtered using structure-based virtual screening (SBVS) to obtain 20 peptides. Of those, three noncompetitive inhibitory peptides (DDQIHIF with an IC50 value of 333.5 µmol·L-1, HDWWKER with an IC50 value of 583.6 µmol·L-1, and THDWWKER with an IC50 value of 1291.8 µmol·L-1) were further investigated based on their favorable pharmacochemical properties and ACE inhibitory activity. Molecular docking studies indicated that the three peptides were entirely enclosed within the ACE protein cavity, improving the overall stability of the complex through interaction forces with the ACE active site. The total free binding energies (ΔGtotal) for DDQIHIF, HDWWKER, and THDWWKER were -21.9 Kcal·mol-1, -71.6 Kcal·mol-1, and -69.1 Kcal·mol-1, respectively. Furthermore, a short-term assay of antihypertensive activity in spontaneously hypertensive rats (SHRs) revealed that HDWWKER could significantly decrease the systolic blood pressure (SBP) of SHRs after intravenous administration. The results showed that based on the better antihypertensive activity of the peptide in SHRs, the feasibility of targeted affinity purification and computer-aided drug discovery (CADD) for the efficient screening and preparation of ACE inhibitory peptide was verified, which provided a new idea of modern drug development method for clinical use.

Anti-Melanogenic Effects of Takifugu flavidus Muscle Hydrolysate in B16F10 Melanoma Cells and Zebrafish.

Abnormal melanogenesis can lead to hyperpigmentation. Tyrosinase (TYR), a key rate-limiting enzyme in melanin production, is an important therapeutic target for these disorders. We investigated the TYR inhibitory activity of hydrolysates extracted from the muscle tissue of Takifugu flavidus (TFMH). We used computer-aided virtual screening to identify a novel peptide that potently inhibited melanin synthesis, simulated its binding mode to TYR, and evaluated functional efficacy in vitro and in vivo. TFMH inhibited the diphenolase activities of mTYR, reducing TYR substrate binding activity and effectively inhibiting melanin synthesis. TFMH indirectly reduced cAMP response element-binding protein phosphorylation in vitro by downregulating melanocortin 1 receptor expression, thereby inhibiting expression of the microphthalmia-associated transcription factor, further decreasing TYR, tyrosinase related protein 1, and dopachrome tautomerase expression and ultimately impeding melanin synthesis. In zebrafish, TFMH significantly reduced black spot formation. TFMH (200 µg/mL) decreased zebrafish TYR activity by 43% and melanin content by 52%. Molecular dynamics simulations over 100 ns revealed that the FGFRSP (T-6) peptide stably binds mushroom TYR via hydrogen bonds and ionic interactions. T-6 (400 µmol/L) reduced melanin content in B16F10 melanoma cells by 71% and TYR activity by 79%. In zebrafish, T-6 (200 µmol/L) inhibited melanin production by 64%. TFMH and T-6 exhibit good potential for the development of natural skin-whitening cosmetic products.

Criticality and Neuromorphic Sensing in a Single Memristor.

Resistive random access memory (RRAM) is an important technology for both data storage and neuromorphic computation, where the dynamics of nanoscale conductive filaments lies at the core of the technology. Here, we analyze the current noise of various silicon-based memristors that involves the creation of a percolation path at the intermediate phase of filament growth. Remarkably, we find that these atomic switching events follow scale-free avalanche dynamics with exponents satisfying the criteria for criticality. We further prove that the switching dynamics are universal and show little dependence on device sizes or material features. Utilizing criticality in memristors, we simulate the functionality of hair cells in auditory sensory systems by observing the frequency selectivity of input stimuli with tunable characteristic frequency. We further demonstrate a single-memristor-based sensing primitive for representation of input stimuli that exceeds the theoretical limits dictated by the Nyquist-Shannon theorem.