Commensal Microbiota Promote Lung Cancer Development via γδ T Cells.

Lung cancer is closely associated with chronic inflammation, but the causes of inflammation and the specific immune mediators have not been fully elucidated. The lung is a mucosal tissue colonized by a diverse bacterial community, and pulmonary infections commonly present in lung cancer patients are linked to clinical outcomes. Here, we provide evidence that local microbiota provoke inflammation associated with lung adenocarcinoma by activating lung-resident γδ T cells. Germ-free or antibiotic-treated mice were significantly protected from lung cancer development induced by Kras mutation and p53 loss. Mechanistically, commensal bacteria stimulated Myd88-dependent IL-1ß and IL-23 production from myeloid cells, inducing proliferation and activation of Vγ6+Vδ1+ γδ T cells that produced IL-17 and other effector molecules to promote inflammation and tumor cell proliferation. Our findings clearly link local microbiota-immune crosstalk to lung tumor development and thereby define key cellular and molecular mediators that may serve as effective targets in lung cancer intervention.

Real-world study of palbociclib combined with endocrine therapy for patients with metastatic breast cancer: A comparison of subsequent treatment patterns and HER2 expression analysis.

BACKGROUND: Cyclin-dependent kinase 4/6 inhibitors combined with endocrine therapy (ET) comprise the standard treatment for patients with hormone receptor-positive and human epidermal growth factor 2 (HER2)-negative metastatic breast cancer. The optimal systematic treatment after progression on palbociclib and the role of HER2 expression among these patients remain unclear. METHODS: The authors retrospectively identified 361 patients who received palbociclib combined with ET. Progression-free survival (PFS) and overall survival (OS) were analyzed based on subsequent treatments and HER2 status (PFSsub and OSsub, respectively). PFS1 and OS1 were calculated from palbociclib administration to disease progression/death and death from any cause, respectively. PFSsub and OSsub were calculated from subsequent treatment initiation. RESULTS: The median PFS1 and OS1 were 10.2 and 39.9 months, respectively. The median PFSsub and OSsub of 111 patients (54.7%) who received chemotherapy were 4.9 months and 20.0 months, respectively, whereas those of 89 patients (43.8%) who received endocrine backbone therapy were 5.9 months and 29.3 months, respectively. Among them, 31 patients (15.3%) who received abemaciclib combined with new ET showed better PFSsub and OSsub (12.2 months and not reached, respectively). The median PFS1 was significantly shorter in the HER2-low subgroup than in the HER2-zero subgroup among patients who received second-line or later palbociclib (6.1 vs. 7.8 months; p = .040) but did not differ among patients who received first-line palbociclib. CONCLUSIONS: Various regimens after palbociclib use were received. An improvement was noted in PFS among patients who received endocrine backbone therapy relative to chemotherapy, which may have been secondary to the receipt of chemotherapy by patients with more aggressive disease. HER2 status was not related to the effect of first-line palbociclib, but it may play a role in later lines.

Persistence of peripheral CD8 + CD28- T cells indicates a favourable outcome and tumour immunity in first-line HER2-positive metastatic breast cancer.

BACKGROUND: The contradictory role of CD8 + CD28- T cells in tumour immunity has been reported, while their biological and clinical significance in HER2-positive metastatic breast cancer (MBC) is still unknown. METHODS: HER2-positive MBC patients with no prior therapy in the metastatic setting were retrospectively recruited at two medical centres. Peripheral CD8 + CD28- T cells (pTCD8+CD28-) were detected at baseline and following therapeutic intervals. Progression-free survival (PFS) was compared according to pTCD8+CD28- levels. The molecular features of pTCD8+CD28- and its correlation with tumour immunity were also investigated. RESULTS: A total of 252 patients were enrolled, and the median follow-up time was 29.6 months. pTCD8+CD28- high at baseline has prolonged PFS compared to pTCD8+CD28- low (P = 0.001). Patients who maintained pTCD8+CD28- high had a longer PFS than those who kept pTCD8+CD28- low (P < 0.001). The enhanced pTCD8+CD28- level also indicates a longer PFS compared to pTCD8+CD28- low (P = 0.025). Here, pTCD8+CD28- was demonstrated as an antigen-experienced effector T cell. Higher IL-2 level (P = 0.034) and lower TGF-ß level (P = 0.016) in the serum and highly infiltrated CD8 + CD28- T cells (P = 0.037) were also connected to pTCD8+CD28- high. CONCLUSIONS: High pTCD8+CD28- level is associated with a favourable tumour immunity and a better PFS of HER2-targeting therapy in MBC patients.

Knockout of lws1 in zebrafish (Danio rerio) reveals its role in regulating feeding and vision-guided behavior.

Long-wave sensitive (LWS) is a G protein-coupled receptor expressed in the retina, and zebrafish is a better model organism for studying vision, but the role of LWS1 in vision-guided behavior of larvae fish has rarely been reported. In this study, we found that zebrafish lws1 and lws2 are tandemly replicated genes, both with six exons, with lws1 being more evolutionarily conserved. The presence of Y277F in the amino acid sequence of lws2 may have contributed to the shift of λmax to green light. We established a lws1 knockout zebrafish model using CRISPR/Cas9 technology. Lws1-/- larvae showed significantly higher levels of feeding and appetite gene (agrp) expression than WT, and significantly lower levels of anorexia gene (pomc, cart) expression. In addition, green light gene compensation was observed in lws1-/- larvae with significantly increased expression levels of rh2-1. The light-dark movement test showed that lws1-/- larvae were more active under light-dark transitions or vibrational stimuli, and the expression of phototransduction-related genes was significantly up-regulated. This study reveals the important role of lws1 gene in the regulation of vision-guided behavior in larvae.

Transcriptome analysis reveals the importance of phototransduction during the first-feeding in mandarin fish (Siniperca chuatsi).

The mandarin fish (Siniperca chuatsi), as a typical freshwater carnivorous fish, has high economic value. Mandarin fish have a peculiar feeding habit of feeding on other live fry during the first-feeding period, while rejecting zooplankton or particulate feed, which may be attributed to the low expression of zooplankton-associated gene sws1 in mandarin fish. The domesticated strain of mandarin fish could feed on Artemia at 3 days post hatching (dph). However, the mechanism of mandarin fish larvae recognize and forage Artemia as food is still unclear. In this study, we employed transcriptional analysis to identify the representative differential pathways between mandarin fish larvae unfed and fed with Artemia at 3 dph. The comparative transcriptome analysis has unveiled a tapestry of genetic expression, highlighting 403 genes that have been up-regulated and 259 that have been down-regulated, all of which constitute the differentially expressed genes (DEGs). KEGG pathway analysis revealed that the number of differentially expressed genes in the photoconductive signaling pathway was the largest. Next, the Vorinostat (suberoylanilide hydroxamic acid, SAHA) was used to assess whether sws1 induced ingestion of Artemia in mandarin fish larvae. We discovered that SAHA-treated larvae had more food intake of Artemia and up-regulated the transcription level of npy, which might have been associated with the up-regulated of sws1 opsin. Additionally, exposure to 0.5 µM SAHA increased the expression of genes involved in phototransduction pathway. These findings would provide insights on the molecular processes involved in mandarin fish larvae feeding on Artemia at the first-feeding stage.

Long-term Outcome Analysis of Pyrotinib in Patients With HER2-Positive Metastatic Breast Cancer and Brain Metastasis: A Real-World Study.

BACKGROUND: Pyrotinib is currently approved for the treatment of HER2-positive advanced breast cancer in China. Data on the overall survival (OS) and efficacy in patients with brain metastasis (BM) remain scarce. This study evaluated the effectiveness of pyrotinib in a real-world setting, especially in patients with BM. METHODS: We reviewed patients with metastatic breast cancer treated with pyrotinib-based therapy between June 2018 and June 2022. Progression-free survival (PFS), OS, objective response rate, and safety were analyzed following the administration of pyrotinib. RESULTS: A total of 239 patients were included. The median PFS in patients who received pyrotinib-based therapy as first-line (15/239), second-line (115/239), or third-or-higher-line (109/239) treatment was 14.00, 9.33, and 8.20 months, respectively, and the median OS was not reached, 29.07 and 22.23 months, respectively. The median PFS in patients who pretreated with trastuzumab (214/239), trastuzumab plus pertuzumab (22/239), lapatinib (68/239), or trastuzumab emtansine (14/239) was 9.33, 6.87, 7.20, and 7.20 months, respectively. In 61 patients with BM, the median PFS was 7.50 months, the median central nervous system (CNS)-PFS was 11.17 months, and the median OS was 21.27 months. Furthermore, 19 patients with concomitant brain radiotherapy tended to achieve a longer OS than 42 patients without radiation (34.17 vs. 20.70 months, P = .112). CONCLUSIONS: Long-term outcomes of pyrotinib-based therapy are promising for patients with HER2-positive metastatic breast cancer in real world and in patients with BM, regardless of the treatment lines and prior anti-HER2 therapies.

Impact of HER2-targeted PET/CT imaging in patients with breast cancer and therapeutic response monitoring.

BACKGROUND: Patients with breast cancer exhibit heterogeneity in the expression of the human epithelial growth factor receptor 2 (HER2). Clinically, re-biopsying recurrent or metastatic lesions presents substantial challenges. This study aimed to evaluate the efficacy of HER2-targeted PET/CT imaging in identifying HER2 expression in breast cancer lesions and monitoring therapeutic responses. PATIENTS AND METHODS: This exploratory analysis used data from a prospective study that included adult patients with breast cancer who underwent both Al18F-NOTA-HER2-BCH and 18F-FDG PET/CT imaging at Beijing Cancer Hospital between June 2020 and July 2023 (NCT04547309). RESULTS: Fifty-nine participants, with a median age of 55 years, were analyzed. Lesions imaged with HER2-targeted PET/CT before anti-HER2 therapy exhibited higher SUVmax values than after therapy in HER2 immunohistochemistry (IHC) 3 + lesions (19.9, 95% CI: 15.7-25.3 vs 9.8, 95% CI: 5.6-14.7; P = .006). A significant positive correlation was observed between SUVmax on HER2-targeted PET/CT and IHC before therapy (P = .034), with higher SUVmax values noted in lesions with positive HER2 pathology compared to those with negative HER2 status (17.9 ±â€…13.2 vs 1.1 ±â€…0.3; P = .007). HER2 expression heterogeneity was confirmed both between primary and metastatic lesions (22.9%) and among different metastatic sites (26.7%) as assessed by HER2-targeted PET/CT. A superior therapeutic response correlated with higher pretreatment SUVmax values. The HER2-targeted PET/CT procedure was well-tolerated by all patients. CONCLUSION: HER2-targeted PET/CT imaging offers a practical, non-invasive, and quantitative approach for assessing HER2 status in breast cancer patients, facilitating the optimization and personalization of therapeutic strategies by oncologists.

OCTA quantitative assessment of exercise-induced variations and recovery in retinal microvasculature of healthy subjects with or without masks.

PURPOSE: To investigate the impact of exercise and mask-wearing on retinal microvasculature using optical coherence tomography angiography (OCTA). METHODS: A total of 30 healthy volunteers were enrolled and tasked with physical exercise to reach 75-80 % maximum heart rates. Swept-source OCTA was performed on the macular region and optic nerve head (ONH) in participants with no mask, surgical mask, or N95 mask at quiescent conditions (Step 1) and 0 min, 10 min, 20 min, and 30 min post-exercise (Steps 2-5, respectively). The functional vessel density (VD), including the superficial and deep plex (SP and DP) in the macular area and the superficial plex (SP), nerve fiber plex, and small vessels in the optic nerve head, were measured. RESULTS: Under quiescent conditions, the functional VD of SP and DP exhibited significant reduction with surgical and N95 masks in the foveal area (P < 0.05). In step 2 (immediately after training) with or without masks, functional VD of SP and nerve fiber both showed significant reduction in the inside disc and peripapillary area, small functional VD of nerve fiber in the ONH showed significant reduction in peripapillary area (P < 0.05). These changes had been recovered in Step 5 (30 min post-exercise) in all groups (no-mask, surgical mask and N95 mask groups) (P > 0.05). CONCLUSIONS: Mask-wearing and physical exercise reduce retinal functional VD in macular and ONH areas. The retinal vasoconstriction induced by exercise tends to recover after rest for approximately 30 min. Our research provides insights into mask-wearing and physical exercise's immediate retinal microvasculature effects, hinting at systemic microvascular changes.

Mechanical Control of Polar Patterns in Wrinkled Thin Films via Flexoelectricity.

Intriguing topological polar structures in oxide nanofilms have drawn growing attention owing to their immense potential applications in nanoscale electronic devices. Here, we report a novel route to mechanically manipulate polar structures via flexoelectricity in wrinkled thin films. Our results present a flexoelectric polar transition from a nonpolar state to uniaxial polar stripes, biaxial meronlike or antimeronlike polar structures, and polar labyrinths by varying wrinkle morphologies. The evolution mechanisms and the outstanding mechanical tunability of these flexoelectric polar patterns were investigated theoretically and numerically. This strategy based on flexoelectricity for generating nontrivial polar structures will no longer rely on the superlattice structure and can be widely applicable to all centrosymmetric or noncentrosymmetric materials, providing a broader range of material and structure candidates for polar topologies.

Spatial and Single-Cell Transcriptomics Reveals the Regional Division of the Spatial Structure of MASH Fibrosis.

OBJECTIVE: To elucidate the regional distribution of metabolic dysfunction-associated steatohepatitis (MASH) fibrosis within the liver and to identify potential therapeutic targets for MASH fibrosis. METHODS: Liver sections from healthy controls, patients with simple steatosis and MASH patients were analysed using spatial transcriptomics integrated with single-cell RNA-seq. RESULTS: Spatial transcriptomics analysis of liver tissues revealed that the fibrotic region (Cluster 9) was primarily distributed in lobules, with some fibrosis also found in the surrounding area. Integration of the single-cell-sequencing data set (GSE189175) showed a greater proportion of inflammatory cells (Kupffer cells and T cells) and myofibroblasts in MASH. Six genes, showing high- or low-specific expression in Cluster 9, namely, ADAMTSL2, PTGDS, S100A6, PPP1R1A, ASS1 and G6PC, were identified in combination with pathology. The average expression levels of ADAMTSL2, PTGDS and S100A6 on the pathological HE staining map were positively correlated with the increase in the degree of fibrosis and aligned strongly with the distribution of fibrosis. ADAMTSL2+ myofibroblasts play a role in TNF signalling pathways and in the production of ECM structural components. Pseudotime analysis indicated that in the early stages of MASH, infiltration by T cells and Kupffer cells triggers a significant inflammatory response. Subsequently, this inflammation leads to the activation of hepatic stellate cells (HSCs), transforming them into myofibroblasts and promoting the development of liver fibrosis. CONCLUSION: This study is the first to characterise lineage-specific changes in gene expression, subpopulation composition, and pseudotime analysis in MASH fibrosis and reveals potential therapeutic targets for this condition.

The function role of HIGD1A in nonalcoholic steatohepatitis from chronic hepatitis B.

BACKGROUND: Accompanied by the growing prevalence of nonalcoholic fatty liver disease (NAFLD), the coexistence of chronic hepatitis B (CHB) and NAFLD has increased. In the context of CHB, there is limited understanding of the factors that influence the development of NASH. METHODS: We enrolled CHB combined NAFLD patients who had liver biopsy and divided them to NASH vs. non-NASH groups. A whole transcriptome chip was used to examine the expression profiles of long noncoding RNAs (lncRNAs) and mRNA in biopsied liver tissues. The function analysis of HIGD1A were performed. We knocked down or overexpressed HIGD1A in HepG2.2.15 cells by transient transfection of siRNA-HIGD1A or pcDNA-HIGD1A. In vivo investigations were conducted using hepatitis B virus (HBV) transgenic mice. RESULTS: In 65 patients with CHB and NAFLD, 28 were patients with NASH, and 37 were those without NASH. After screening 582 differentially expressed mRNAs, GO analysis revealed differentially expressed mRNAs acting on nicotinamide adenine dinucleotide phosphate (NADPH), which influenced redox enzyme activity. KEGG analysis also shown that they were involved in the NAFLD signaling pathway. The function analysis revealed that HIGD1A was associated with the mitochondrion. Then, both in vivo and in vitro CHB model, HIGD1A was significantly higher in the NASH group than in the non-NASH group. HIGD1A knockdown impaired mitochondrial transmembrane potential and induced cell apoptosis in HepG2.2.15 cells added oleic acid and palmitate. On the contrary, hepatic HIGD1A overexpression ameliorated free fatty acids-induced apoptosis and oxidative stress. Furthermore, HIGD1A reduced reactive oxygen species (ROS) level by increasing glutathione (GSH) expression, but Adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK)/Acetyl-CoA carboxylase (ACC) pathway was not involved. CONCLUSION: Both in vivo and in vitro CHB model, an upward trend of HIGD1A was observed in the NASH-related inflammatory response. HIGDIA played a protective role in cells against oxidative stress. Our data suggested that HIGD1A may be a positive regulator of NASH within the CHB context.

Deep-learning-based radiomics of intratumoral and peritumoral MRI images to predict the pathological features of adjuvant radiotherapy in early-stage cervical squamous cell carcinoma.

BACKGROUND: Surgery combined with radiotherapy substantially escalates the likelihood of encountering complications in early-stage cervical squamous cell carcinoma(ESCSCC). We aimed to investigate the feasibility of Deep-learning-based radiomics of intratumoral and peritumoral MRI images to predict the pathological features of adjuvant radiotherapy in ESCSCC and minimize the occurrence of adverse events associated with the treatment. METHODS: A dataset comprising MR images was obtained from 289 patients who underwent radical hysterectomy and pelvic lymph node dissection between January 2019 and April 2022. The dataset was randomly divided into two cohorts in a 4:1 ratio.The postoperative radiotherapy options were evaluated according to the Peter/Sedlis standard. We extracted clinical features, as well as intratumoral and peritumoral radiomic features, using the least absolute shrinkage and selection operator (LASSO) regression. We constructed the Clinical Signature (Clinic_Sig), Radiomics Signature (Rad_Sig) and the Deep Transformer Learning Signature (DTL_Sig). Additionally, we fused the Rad_Sig with the DTL_Sig to create the Deep Learning Radiomic Signature (DLR_Sig). We evaluated the prediction performance of the models using the Area Under the Curve (AUC), calibration curve, and Decision Curve Analysis (DCA). RESULTS: The DLR_Sig showed a high level of accuracy and predictive capability, as demonstrated by the area under the curve (AUC) of 0.98(95% CI: 0.97-0.99) for the training cohort and 0.79(95% CI: 0.67-0.90) for the test cohort. In addition, the Hosmer-Lemeshow test, which provided p-values of 0.87 for the training cohort and 0.15 for the test cohort, respectively, indicated a good fit. DeLong test showed that the predictive effectiveness of DLR_Sig was significantly better than that of the Clinic_Sig(P < 0.05 both the training and test cohorts). The calibration plot of DLR_Sig indicated excellent consistency between the actual and predicted probabilities, while the DCA curve demonstrating greater clinical utility for predicting the pathological features for adjuvant radiotherapy. CONCLUSION: DLR_Sig based on intratumoral and peritumoral MRI images has the potential to preoperatively predict the pathological features of adjuvant radiotherapy in early-stage cervical squamous cell carcinoma (ESCSCC).

HER2 status results in an unstable switch from primary to recurrent breast cancer.

Accurately distinguishing HER2-2+ tumors from HER2-0/1+ tumors via immunohistochemistry (IHC) is still very challenging. HER2 IHC 2+ is considered to indicate moderate expression and is easier to distinguish, with more reliable results in previous and current clinical practice. We focused on HER2-2+ patients and evaluated the switch in HER2 status between primary and paired recurrent disease patients to evaluate the discordance of HER2-2+ expression. We included patients who were HER2-2+ of primary or rebiopsy tumor samples, to evaluate the evolution of HER2-2+ expression. In the cohort with a total of 159 patients with HER2-2+ expression in either primary tumor or locoregional/distant metastasis samples, 44.0% had HER2-2+ in primary tumor and 88.8% in recurrent disease. Among patients with primary and recurrent HER2-2+ breast cancers, 18.5% and 15.2% of the patients, respectively, had HER2 gene amplification via ISH. The overall rate of discordance in HER2 IHC results was 67.1%. Among primary HER2-2+ patients, 74.6% were maintained in the HER2-2+ cohort at the recurrence. The discordance was mostly driven by patients switching from HER2-2+ to HER2-1+ (64.7%). Among HER2-2+ recurrent patients, discordance in the IHC results was mostly driven by switching from HER2-0 to HER2-2+ (47.1%). When HER2-low was added to the analysis, the overall rate of HER2 discordance was 40.4%. The proportion of patients with discordant HER2 expression was significantly greater among HR-positive patients than negative patients (44.1% vs. 21.7%, p=0.062). HER2 expression in primary and recurrent breast cancer samples was highly unstable. Discordance was more frequently observed in the HR-positive population.

L-Glutamate Regulates Npy via the mGluR4-Ca2+-ERK1/2 Signaling Pathway in Mandarin Fish (Siniperca chuatsi).

Metabotropic glutamate receptor 4 (mGluR4) is widely regarded as an umami receptor activated by L-glutamate to exert essential functions. Numerous studies have shown that umami receptors participate in food intake regulation. However, little is known about mGluR4's role in mediating food ingestion and its possible molecular mechanism. Mandarin fish, a typical carnivorous fish, is sensitive to umami substances and is a promising vertebrate model organism for studying the umami receptor. In this study, we identified the mGluR4 gene and conducted evolutionary analyses from diverse fish species with different feeding habits. mGluR4 of mandarin fish was cloned and functionally expressed to investigate the effects of L-glutamate on mGluR4. We further explored whether the signal pathway mGluR4-Ca2+-ERK1/2 participates in the process in mandarin fish brain cells. The results suggest that L-glutamate could regulate Neuropeptide Y (Npy) via the mGluR4-Ca2+-ERK1/2 signaling pathway in mandarin fish. Our findings unveil the role of mGluR4 in feeding decisions and its possible molecular mechanisms in carnivorous fishes.

Diterpenoids with Potent Anti-Psoriasis Activity from Euphorbia helioscopia L.

Psoriasis, an immune-mediated inflammatory skin disorder, seriously affects the quality of life of nearly four percent of the world population. Euphorbia helioscopia L. is the monarch constituent of Chinese ZeQi powder preparation for psoriasis, so it is necessary to illustrate its active ingredients. Thus, twenty-three diterpenoids, including seven new ones, were isolated from the whole herb of E. helioscopia L. Compounds 1 and 2, each featuring a 2,3-dicarboxylic functionality, are the first examples in the ent-2,3-sceo-atisane or the ent-2,3-sceo-abietane family. Extensive spectroscopic analysis (1D, 2D NMR, and HRMS data) and computational methods were used to confirm their structures and absolute configurations. According to the previous study and NMR data from the jatropha diterpenes obtained in this study, some efficient 1H NMR spectroscopic rules for assigning the relative configurations of 3α-benzyloxy-jatroph-11E-ene and 7,8-seco-3α-benzyloxy-jatropha-11E-ene were summarized. Moreover, the hyperproliferation of T cells and keratinocytes is considered a key pathophysiology of psoriasis. Anti-proliferative activities against induced T/B lymphocytes and HaCaT cells were tested, and IC50 values of some compounds ranged from 6.7 to 31.5 µM. Compounds 7 and 11 reduced the secretions of IFN-γ and IL-2 significantly. Further immunofluorescence experiments and a docking study with NF-κB P65 showed that compound 13 interfered with the proliferation of HaCaT cells by inhibiting the NF-κB P65 phosphorylation at the protein level.

Dietary supplementation of VA enhances growth, feed utilization, glucose and lipid metabolism, appetite, and antioxidant capacity of Chinese perch (Siniperca chuatsi).

The aim of this study was to investigate the effect of dietary vitamin A on juvenile Chinese perch (Siniperca chuatsi). Chinese perch were fed with five experimental diets containing 0, 20, 40, 60, and 80 mg VA·kg-1 for 8 weeks. Results showed that dietary vitamin A significantly influenced the fish's growth, feed utilization, glucose and lipid metabolism, appetite, and antioxidant capacity. Vitamin A-supplemented groups had higher weight gain rate (WGR) and specific growth rate (SGR) compared to the control group. Feed conversion ratio (FCR) was also lower in the vitamin A-supplemented groups. Dietary vitamin A had no significant effect on the survival rate (SR). Compared to the control group, fish fed with vitamin A had increased feed intake (FI), and the expression of appetite-promoting genes (npy and agrp) was significantly higher in the 40 mg VA·kg-1 group. Vitamin A also enhanced the utilization of dietary protein by Chinese perch. The serum glucose content of the fish fed with 40 mg VA·kg-1 diet was significantly higher than that of the control group and 20 mg VA·kg-1 diet, indicating that the promoting effect of VA on gluconeogenesis was greater than that on glycolysis. Additionally, dietary vitamin A increased the expression of lipid metabolism-related genes (hl and fas) and antioxidant genes (nrf2 and gpx) in the fish. These results suggest that the optimal vitamin A requirement of juvenile Chinese perch bream was estimated to be 37.32 mg VA·kg-1 based on broken-line regression analysis of WGR. In conclusion, this study provides valuable insights into the potential benefits of dietary vitamin A on the growth, metabolism, and antioxidant capacity of Chinese perch.

Addition of α-ketoglutaric acid (AKG) reduces deamination in Chinese perch (Siniperca chuatsi) fed with fermented soybean meal as a substitute for fishmeal.

To alleviate amino acid imbalances in fermented soybean meal as a replacement for fishmeal feeds, this study evaluated the effects of adding lysine (Lys), methionine (Met), and α-ketoglutaric acid (AKG) to fermented soybean meals for Chinese perch. Chinese perch (34 ± 3 g) were fed five diets for 66 days (fishmeal as the protein source of the basal diet [FM]; fermented soybean meal as a substitute for 30% fishmeal in the soybean meal diet [FSM]; addition of crystalline Lys and Met [AA]; addition of α-ketoglutaric acid [AKG]; and simultaneous addition of crystalline Lys, Met, and AKG [BA] to the soybean meal diet). At the end of the feeding trial, the FSM group had the highest feeding rate and the lowest weight gain rate among all the groups. The FM group had the highest protein retention and the lowest feed efficiency among the groups. The mRNA transcription level of genes related to the AMP-activating protein (AMPK) signaling pathway and amino acid response (AAR) signaling pathway (lkb1, atf4, and gcn2) were highest in the AA group (P < 0.05) but lower in the AKG and BA groups. In the AKG group, the mRNA transcription level of the gluconeogenesis pathway-related gene (pepck and g6pase) was significantly higher than that in the other four groups, but the mRNA transcription level of genes related to amino acid catabolism (gdh and ampd) was lower. Among all the groups, the FSM group had the lowest mRNA transcription level of genes associated with the mammalian target of rapamycin (mTOR) signaling pathway (mtor and s6k). These findings imply that the feeding rate of Chinese perch in the fermented soybean meal group was the highest, but the protein retention was the lowest, while the addition of Lys, Met, and AKG improved protein retention. In conclusion, the addition of AKG to fermented soybean meal as a fishmeal substitute reduced amino acid deamination, enhanced gluconeogenesis, and increased protein deposition, which contributed to the growth of Chinese perch, alleviated amino acid imbalances, and improved the feed utilization of Chinese perch.

Dissolved oxygen and ammonia affect ammonia production via GDH/AMPK signaling pathway and alter flesh quality in Chinese perch (Siniperca chuatsi).

The dissolved oxygen (DO) and ammonia are crucial to the growth of Chinese perch (Siniperca chuatsi). Information on the effects of DO and total ammonia nitrogen (TAN) in regulating ammonia nitrogen excretion and flesh quality in Chinese perch is scanty. This study aimed to evaluate the effects of dissolved DO at oxygen levels of 3 mg/L and 9 mg/L, as well as the TAN concentrations of 0.3 mg/L and 0.9 mg/L on ammonia excretion and flesh quality. Results showed that the ammonia contents in plasma, muscle, and liver of the 9 mg/L DO group were significantly higher than those of the 3 mg/L DO group (P < 0.05). However, the expression of AMPK-related signaling pathway genes (gdh, lkb1, and ampd) and flesh quality indicators (gumminess, chewiness, hardness) in the 9 mg/L DO group were significantly lower than those in the 3 mg/L DO group. Under long-term exposure to 0.9 mg/L TAN, the ammonia contents in plasma and gill filaments, as well as muscle flesh quality (resilience, gumminess, chewiness, cohesiveness), were significantly lower than those in the 0.3 mg/L TAN group (P < 0.05). However, the activities of GDH and AMPD enzymes in the 0.9 mg/L TAN group were significantly higher than those in the 0.3 mg/L TAN group. In summary, when fish are exposed to 3 mg/L DO and 0.9 mg/L TAN in the environment for a long time, their amino acids are used for transamination and deamination, resulting in insufficient energy supply for Chinese perch, whereas 9 mg/L DO and 0.9 mg/L TAN caused deterioration of the flesh quality.

Regulatory effect of NK homeobox 1 (NKX2.1) on melanocortin 4 receptor (Mc4r) promoter in Mandarin fish.

The melanocortin 4 receptor (MC4R) is a G protein-coupled transporter that mediates the regulation of thyroid hormones and leptin on energy balance and food intake. However, the mechanisms of transcriptional regulation of Mc4r by thyroid hormone and leptin in fish have been rarely reported. The messenger RNA expression of Mc4r gene was significantly higher in brain than those in other tissues of mandarin fish. We analyzed the structure and function of a 2029 bp sequence of Mc4r promoter. Meanwhile, overexpression of NKX2.1 and incubation with leptin significantly increased Mc4r promoter activity, but triiodothyronine showed the opposite effect. In addition, mutations in the NKX2.1 binding site abolished not only the activation of Mc4r promoter activity by leptin but also the inhibitory effect of thyroid hormones on Mc4r promoter activity. In summary, these results suggested that thyroid hormones and leptin might regulate the transcriptional expression of Mc4r through NKX2.1.

Role of phosphoenolpyruvate carboxykinase 1 (pck1) in mediating nutrient metabolism in zebrafish.

Carbohydrates are the most economical source of energy in fish feeds, but most fish have limited ability to utilize carbohydrates. It has been reported that phosphoenolpyruvate carboxykinase 1 (pck1) is involved in carbohydrate metabolism, lipid metabolism, and other metabolic processes. However, direct evidence is lacking to fully understand the relationship between pck1 and glucose and lipid metabolism. Here, we generated a pck1 knockout zebrafish by CRISPR/cas9 system, and a high-carbohydrate diet was provided to 60 days post-fertilization (dpf) for 8 weeks. We found that pck1-deficient zebrafish displayed decreased plasma glucose, elevated mRNA levels of glycolysis-related genes (gck, pfk, pk), and reduced the transcriptional levels of gluconeogenic genes (pck1, fbp1a) in liver. We also found decreased triglyceride, total cholesterol, and lipid accumulation and in pck1-/- zebrafish, along with downregulation of genes for lipolysis (acaca) and lipogenesis (cpt1). In addition, the observation of HE staining revealed that the total muscle area of pck1-/- was substantially less than that of WT zebrafish and real-time PCR suggested that GH/IGF-1 signaling (ulk2, stat1b) may be suppressed in pck1-deficient fish. Taken together, these findings suggested that pck1 may play an important role in the high-carbohydrate diet utilization of fish and significantly affected lipid metabolism and protein synthesis in zebrafish. pck1 knockout mutant line could facilitate a further mechanism study of pck1-associated metabolic regulation and provide new information for improving carbohydrate utilization traits.