Increasing evidence indicates that brown adipose tissue (BAT) transplantation enhances whole-body energy metabolism in a mouse model of diet-induced obesity. However, it remains unclear whether BAT also has such beneficial effects on genetically obese mice. To address this issue, we transplanted BAT from C57/BL6 mice into the dorsal subcutaneous region of age- and sex-matched leptin deficient Ob/Ob mice. Interestingly, BAT transplantation led to a significant reduction of body weight gain with increased oxygen consumption and decreased total body fat mass, resulting in improvement of insulin resistance and liver steatosis. In addition, BAT transplantation increased the level of circulating adiponectin, whereas it reduced the levels of circulating free T3 and T4, which regulate thyroid hormone sensitivity in peripheral tissues. BAT transplantation also increased β3-adrenergic receptor and fatty acid oxidation related gene expression in subcutaneous and epididymal (EP) white adipose tissue. Accordingly, BAT transplantation increased whole-body thermogenesis. Taken together our results demonstrate that BAT transplantation may reduce obesity and its related diseases by activating endogenous BAT.

Broad-spectrum drugs appear to be more promising for the treatment of acute ischemic stroke. In our previous work, a new ligustrazine derivative (3,5,6-trimethylpyrazin-2-yl) methyl 3-methoxy-4-[(3,5,6-trimethylpyrazin-2-yl)methoxy]benzoate (T-VA) showed neuroprotective effect on injured PC12 cells (EC50 = 4.249 µM). In the current study, we show that this beneficial effect was due to the modulation of nuclear transcription factor-κB/p65 (NF-κB/p65) and cyclooxygenase-2 (COX-2) expressions. We also show that T-VA exhibited neuroprotective effect in a rat model of ischemic stroke with concomitant improvement of motor functions. We propose that the protective effect observed in vivo is owing to increased vascular endothelial growth factor (VEGF) expression, decreased oxidative stress, and up-regulation of Ca(2+)-Mg(2+) ATP enzyme activity. Altogether, our results warrant further studies on the utility of T-VA for the potential treatment of ischemic brain injuries, such as stroke.

In this study, a bioinformatics analysis and luciferase reporter assay revealed that microRNA-141 could silence the expression of lncRNA-HOTAIR by binding to specific sites on lncRNA-HOTAIR. We used superparamagnetic iron oxide nanoparticles (SPIONs) to mediate the high expression of microRNA-141 (SPIONs@miR-141) in human amniotic epithelial stem cells (HuAESCs), which was followed by the induction of the differentiation of HuAESCs into dopaminergic neuron-like cells (iDNLCs). qPCR, western blot, immunofluorescence staining and HPLC all suggested that SPION-mediated overexpression of miR-141 could promote an increased expression of brain-derived neurotrophic factor (BDNF), DAT and 5-TH in HuAESC-derived iDNLCs. The RIP and ChIP assay also showed that overexpression of miR-141 could significantly inhibit the recruitment and binding of lncRNA-HOTAIR to EZH2 on BDNF gene promoter. cDNA microarray analysis revealed that the expression levels of 190 genes were much higher in iDNLCs than in HuAESCs. Finally, a protein interaction network analysis and identification showed that in the iDNLC group with SPIONs@miR-141, factors that interact with BDNF, such as FGF8, SHH, NTRK3 and CREB1, all showed significantly higher expression levels compared with those in the SPIONs@miR-Mut. Therefore, this study confirmed that the highly efficient expression of microRNA-141 mediated by SPIONs could improve the efficiency of HuAESCs differentiation into dopaminergic neuron-like cells.

Stress-induced hyperglycemia is a fundamental adaptive response that mobilizes energy stores in response to threats. Here, our examination of the contributions of the central catecholaminergic (CA) neuronal system to this adaptive response revealed that CA neurons in the ventrolateral medulla (VLM) control stress-induced hyperglycemia. Ablation of VLM CA neurons abolished the hyperglycemic response to both physical and psychological stress, whereas chemogenetic activation of these neurons was sufficient to induce hyperglycemia. We further found that CA neurons in the rostral VLM, but not those in the caudal VLM, cause hyperglycemia via descending projections to the spinal cord. Monosynaptic tracing experiments showed that VLM CA neurons receive direct inputs from multiple stress-responsive brain areas. Optogenetic studies identified an excitatory PVN-VLM circuit that induces hyperglycemia. This study establishes the central role of VLM CA neurons in stress-induced hyperglycemia and substantially expands our understanding of the central mechanism that controls glucose metabolism.

Recent field data showed that lumbar spine fractures occurred more frequently in late model vehicles than early ones in frontal crashes. However, the lumbar spine designs of the current crash test dummies are not accurate in human anatomy and have not been validated against any human/cadaver impact responses. In addition, the lumbar spines of finite element (FE) human models, including GHBMC and THUMS, have never been validated previously against cadaver tests. Therefore, this study developed a detailed FE lumbar spine model and validated it against cadaveric tests. To investigate the mechanism of lumbar spine injury in frontal crashes, effects of changing the coefficient of friction (COF), impact velocity, cushion thickness and stiffness, and cushion angle on the risk of lumbar spine injuries were analyzed based on a Taguchi array of design of experiments. The results showed that impact velocity is the most important factor in determining the risk of lumbar spine fracture (P = 0.009). After controlling the impact velocity, increases in the cushion thickness can effectively reduce the risk of lumbar spine fracture (P = 0.039).

Spermatogonial stem cell (SSC) transplantation is an alternative reproductive method to achieve conservation and production of elite animals in livestock production. Creating a recipient animal without endogenous germ cells is important for effective SSC transplantation. However, natural mutants with depletion of SSCs are difficult to obtain, and drug ablation of endogenous germ cells is arduous to perform for practical use. In this study, we used mouse models to study the preparation of recipients with congenital germ cell ablation. We knocked out (KO) Ets-variant gene 5 (Etv5) in mice using the CRISPR/Cas9 system. The testicular weight of Etv5-/- mice was significantly lower than that of wild-type (WT) mice. The germ cell layer of the seminiferous tubules gradually receded with age in Etv5-/- mice. At 12 weeks of age, the tubules of Etv5-/- mice lacked almost all spermatogenic cells with a Sertoli cell-only phenotype, and sperm were completely absent in the epididymis. We subsequently transplanted allogeneic SSCs with enhanced green fluorescent protein (EGFP) into 3- (immature) or 7-week-old (mature) Etv5-/- mice. Partial restoration of germ cell layers in the seminiferous tubules and spermatogenesis was observed in all immature testes but not in mature adult testes at 2 months post-transplantation. The presence of heterologous genes Etv5 and EGFP in recipient testicular tissue and epididymal sperm by PCR indicated that sperm originated from the transplanted donor cells. Our study demonstrates that, although Etv5-/- mice could accommodate and support foreign germ cell transplantation, this process occurs in a quite low efficiency to support a full spermatogenesis of transplanted SSCs. However, using Etv5-/- mice as a recipient model for SSC transplantation is feasible, and still needs further investigation to establish an optimized transplantation process.

Clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (Cas9) is a precise genome manipulating tool that can produce targeted gene mutations in various cells and organisms. Although CRISPR/Cas9 can efficiently generate gene knockout, the gene knock-in (KI) efficiency mediated by homology-directed repair remains low, especially for large fragment integration. In this study, we established an efficient method for the CRISPR/Cas9-mediated integration of large transgene cassette, which carries salivary gland-expressed multiple digestion enzymes (≈ 20 kbp) in CEP112 locus in pig fetal fibroblasts (PFFs). Our results showed that using an optimal homology donor with a short and a long arm yielded the best CRISPR/Cas9-mediated KI efficiency in CEP112 locus, and the targeting efficiency in CEP112 locus was higher than in ROSA26 locus. The CEP112 KI cell lines were used as nuclear donors for somatic cell nuclear transfer to create genetically modified pigs. We found that KI pig (705) successfully expressed three microbial enzymes (β-glucanase, xylanase, and phytase) in salivary gland. This finding suggested that the CEP112 locus supports exogenous gene expression by a tissue-specific promoter. In summary, we successfully targeted CEP112 locus in pigs by using our optimal homology arm system and established a modified pig model for foreign digestion enzyme expression in the saliva.

As the most common malignant disease in childhood, children acute lymphoblastic leukemia (ALL) is a heterogeneous disease caused by the accumulated genetic alterations. Long non-coding RNAs (lncRNAs) are reported as critical regulators in diseases. GEPIA database indicated that long intergenic non-protein coding RNA 221 (LINC00221) was conspicuously down-regulated in acute myeloid leukemia. However, its expression pattern in ALL has not been revealed. This work was carried out to study the role of LINC00221 in ALL cells. Quantitative real-time PCR (qRT-PCR) quantified LINC00221 expression in ALL cells. The function of LINC00221 in ALL was determined by ki-67 immunofluorescence staining, EdU, TUNEL, JC-1, and caspase-3/8/9 activity assays. RNA pull down and Ago2-RNA immunoprecipitation (RIP) assays investigated the interaction between miR-152-3p and LINC00221 or ATPase sarcoplasmic/endoplasmic reticulum Ca2+ transporting 2 (ATP2A2). Our study revealed the low expression of LINC00221 in ALL cells. Subsequently, LINC00221 was verified to bind with miR-152-3p. Moreover, functional assays pointed out that LINC00221 overexpression posed anti-proliferation and pro-apoptosis effects in ALL cells, and these effects could be separately reversed by miR-152-3p up-regulation. Afterward, LINC00221 was revealed to regulate ATP2A2 expression via sponging miR-152-3p. Additionally, ATP2A2 was verified to involve in regulating LINC00221-mediated ALL cell proliferation and apoptosis. In conclusion, LINC00221 suppressed ALL cell proliferation and boosted ALL cell apoptosis via sponging miR-152-3p to up-regulate ATP2A2.

Shrimp surimi is widely acknowledged as a value-added shrimp product due to its delicious taste, rich flavor, and nutrition. However, the refrigerated shrimp surimi is prone to deterioration due to rapid microbial growth during storage. The present study sought to assess the effects of curcumin-mediated sono/photodynamic treatment on bacterial spoilage and shrimp surimi quality stored at 4 °C. The total viable count (TVC), microbiota composition, and quality parameters, including the total volatile basic nitrogen (TVB-N), thiobarbituric acid reactive substance (TBARs), and pH were investigated. The results showed that the spoilage bacteria in shrimp surimi rapidly increased with a surge on day 2 during refrigeration storage. The Psychrobacter and Brochothrix were identified as the Specific Spoilage Organisms (SSOs), which were also positively correlated with TVB-N and TBARs. The results further elucidated that the sono/photodynamic treatment could significantly inhibit the growth of SSOs on the surface and interior of shrimp surimi and delay shrimp surimi quality deterioration. In conclusion, the sono/photodynamic treatment as a non-thermal sterilization method could be a reliable and potential method for inactivating spoilage microorganisms and preserving shrimp surimi quality.

Klebsiella pneumoniae is capable of acquiring various exogenous genetic elements and subsequently conferring high antimicrobial resistance. Recently, a plasmid-mediated RND family multidrug efflux pump gene cluster, tmexCD1-toprJ1, was discovered in K. pneumoniae. In this study, we analyzed tigecycline-resistant K. pneumoniae isolates from patients from surveillance from 2017 to 2021. In addition to phenotype detection, including growth curves, plasmid transferability and stability, hypermucoviscosity, biofilm formation, and serum survival, by whole-genome sequencing, we analyzed the phylogenetic relationships of the isolates harboring tmexCD1-toprJ1 and discovered the composition of plasmids carrying tmexCD1-toprJ1. In total, we discovered that 12 tigecycline-resistant isolates from 5 patients possessed tmexCD1-toprJ1, designated sequence type 22 (ST22) and ST3691. An ST11 isolate harbored a partial tmexD1, and a complete toprJ1 (tmexC1 was lost) was tigecycline sensitive. All the ST22 tigecycline-resistant isolates coharbored tmexCD1-toprJ1, blaNDM-1, and blaKPC-2. tmexCD1-toprJ1 was encoded by a novel IncU plasmid in ST22 and an IncFIB/HI1B plasmid in ST3691, which presented differences in mobility and stability. Interestingly, isolates from the same patients presented heteroresistance to tigecycline, not only among isolates from different specimens but also those from the same sample, which might be attributed to the differential expression of tmexCD1-toprJ1 due to the dynamic genetic heterogeneity caused by relocating tmexCD1-toprJ1 close to the replication origin of plasmid. Here, we reported the emergence of K. pneumoniae isolates coharboring tmexCD1-toprJ1, blaNDM-1, and blaKPC-2. The results highlight the impact of in vivo genetic heterogeneity of tmexCD1-toprJ1-carrying elements on the in vivo variation of tigecycline resistance, which might have notable influences on antimicrobial treatment. IMPORTANCE Pandrug-resistant (PDR) Klebsiella pneumoniae poses a great challenge to public health, and tigecycline is an essential choice for antimicrobial treatment. In this study, we reported the emergence of PDR K. pneumoniae coharboring tmexCD1-toprJ1, blaNDM-1, and blaKPC-2, which belongs to ST22 and ST3691. By whole-genome analysis, we reconstructed the evolutionary map of the ST22 ancestor to become the PDR superbug by acquiring multiple genetic elements encoding tmexCD1-toprJ1 or blaNDM-1. Importantly, the genetic contexts of tmexCD1-toprJ1 among the ST22 isolates are different and present with various mobilities and stabilities. Furthermore, we also discovered the heterogeneity of tigecycline resistance during long-term infection of ST22, which might be attributed to the differential expression of tmexCD1-toprJ1 due to the dynamic genetic heterogeneity caused by relocating tmexCD1-toprJ1 close to the replication origin of plasmid. This study tracks the inter- and intrahost microevolution of the superbug PDR K. pneumoniae and highlights the importance of timely monitoring of the variation of pathogens during antimicrobial treatment.

Phenotypes associated with metabolism and water retention are thought to be key to the adaptation of desert species. However, knowledge on the genetic changes and selective regimes on the similar and divergent ways to desert adaptation in sympatric and phylogenetically close desert organisms remains limited. Here, we generate a chromosome level genome assembly for Northern three-toed jerboa (Dipus sagitta) and three other high-quality genome assemblies for Siberian jerboa (Orientallactaga sibirica), Midday jird (Meriones meridianus), and Desert hamster (Phodopus roborovskii). Genomic analyses unveil that desert adaptation of the four species mainly result from similar metabolic pathways, such as arachidonic acid metabolism, thermogenesis, oxidative phosphorylation, insulin related pathway, DNA repair and protein synthesis and degradation. However, the specific evolved genes in the same adaptative molecular pathway often differ in the four species. We also reveal similar niche selection but different demographic histories and sensitivity to climate changes, which may be related to the diversified genomic adaptative features. In addition, our study suggests that nocturnal rodents have evolved some specific adaptative mechanism to desert environments compared to large desert animals. Our genomic resources will provide an important foundation for further research on desert genetic adaptations.

This study aimed to identify the clinical outcomes, microbiological features and risk factors for difficult-to-treat resistance (DTR) Klebsiella pneumoniae (Kp) infection.

The type V-F CRISPR-Cas12f system is a strong candidate for therapeutic applications due to the compact size of the Cas12f proteins. In this work, we identify six uncharacterized Cas12f1 proteins with nuclease activity in mammalian cells from assembled bacterial genomes. Among them, OsCas12f1 (433 aa) from Oscillibacter sp. and RhCas12f1 (415 aa) from Ruminiclostridium herbifermentans, which respectively target 5' T-rich Protospacer Adjacent Motifs (PAMs) and 5' C-rich PAMs, show the highest editing activity. Through protein and sgRNA engineering, we generate enhanced OsCas12f1 (enOsCas12f1) and enRhCas12f1 variants, with 5'-TTN and 5'-CCD (D = not C) PAMs respectively, exhibiting much higher editing efficiency and broader PAMs, compared with the engineered variant Un1Cas12f1 (Un1Cas12f1_ge4.1). Furthermore, by fusing the destabilized domain with enOsCas12f1, we generate inducible-enOsCas12f1 and demonstate its activity in vivo by single adeno-associated virus delivery. Finally, dead enOsCas12f1-based epigenetic editing and gene activation can also be achieved in mammalian cells. This study thus provides compact gene editing tools for basic research with remarkable promise for therapeutic applications.

Fucoxanthin, a natural carotenoid abundant in edible brown seaweeds, has been shown to possess anti-cancer, anti-oxidant, anti-obesity and anti-diabetic effects. In this study, we report for the first time that fucoxanthin effectively protects against scopolamine-induced cognitive impairments in mice. In addition, fucoxanthin significantly reversed the scopolamine-induced increase of acetylcholinesterase (AChE) activity and decreased both choline acetyltransferase activity and brain-derived neurotrophic factor (BDNF) expression. Using an in vitro AChE activity assay, we discovered that fucoxanthin directly inhibits AChE with an IC50 value of 81.2 μM. Molecular docking analysis suggests that fucoxanthin likely interacts with the peripheral anionic site within AChE, which is in accordance with enzymatic activity results showing that fucoxanthin inhibits AChE in a non-competitive manner. Based on our current findings, we anticipate that fucoxanthin might exhibit great therapeutic efficacy for the treatment of Alzheimer's disease by acting on multiple targets, including inhibiting AChE and increasing BDNF expression.

Multiple sclerosis (MS) is an inflammatory demyelinating disease of CNS. Astragalus polysaccharides (APS), the main active extract from astragalus membranaceus which is a kind of traditional Chinese medicinal herb, is associated with a variety of immunomodulatory activities. We have evaluated the therapeutic effects of APS in the animal model of MS, experimental autoimmune encephalomyelitis (EAE). It was found that APS could effectively alleviate EAE through inhibiting MOG35-55-specific T cell proliferation and reducing the expression of proinflammatory cytokines, which is mediated by up-regulating the expression of PD-1/PD-Ls signaling pathway. Our results demonstrated that EAE could be suppressed significantly by APS administration. It indicated that APS might be a potential of developing innovative drug for the therapy of MS.

In pig production, inefficient feed digestion causes excessive nutrients such as phosphorus and nitrogen to be released to the environment. To address the issue of environmental emissions, we established transgenic pigs harboring a single-copy quad-cistronic transgene and simultaneously expressing three microbial enzymes, β-glucanase, xylanase, and phytase in the salivary glands. All the transgenic enzymes were successfully expressed, and the digestion of non-starch polysaccharides (NSPs) and phytate in the feedstuff was enhanced. Fecal nitrogen and phosphorus outputs in the transgenic pigs were reduced by 23.2-45.8%, and growth rate improved by 23.0% (gilts) and 24.4% (boars) compared with that of age-matched wild-type littermates under the same dietary treatment. The transgenic pigs showed an 11.5-14.5% improvement in feed conversion rate compared with the wild-type pigs. These findings indicate that the transgenic pigs are promising resources for improving feed efficiency and reducing environmental impact.

Poor initial stability at the first four weeks after surgery is becoming the major causes for metal implant failure. Previous attempts neglected the control release of insulin for the bone regeneration among nondiabetic subjects. The major reason may lie in the adverse effects, such as attenuated bone formation, hypoglycemia or hyperinsulinemia, that caused by the excessive insulin. Thus, spatiotemporal release of insulin may serve as the promising strategy. To address this, through solvent extraction (EMS), solvent evaporation (SMS) and cosolvent methods (CMS), we prepared three types of PLGA microspheres with various internal structures, but similar size distribution. The effects of the preparation methods on the properties of the microspheres, such as their release behavior, degradation of molecular weight, and structural evolution, were investigated. Human bone marrow mesenchymal stromal cells (BMSCs) and rabbit implant models were used to test the bioactivity of the microspheres in vitro and in vivo, respectively. The result demonstrated that these three preparation methods did not influence the polymer degradation but instead affected the internal structural evolution, which plays a crucial role in the release behavior, osteogenesis and peri-implant bone regeneration. Compared with EMS and CMS microspheres, SMS microspheres exhibited a relatively steady release rate in the first four weeks, which evidently stimulated the osteogenic differentiation of the stem cells and peri-implant bone regeneration. Meanwhile, SMS microspheres significantly enhanced the stability of the implant at Week 4, which is promising to reduce early failure rate of the implant without inducing adverse effects on the serum biochemical indices.

The salivary glands of animals have great potential to act as powerful bioreactors to produce human therapeutic proteins. Human nerve growth factor (hNGF) is an important pharmaceutical protein that is clinically effective in the treatment of many human neuronal and non-neuronal diseases. In this study, we generated 18 transgenic (TG) founder mice each carrying a salivary gland specific promoter-driven hNGF transgene. A TG mouse line secreting high levels of hNGF protein in its saliva (1.36 μg/mL) was selected. hNGF protein was successfully purified from the saliva of these TG mice and its identity was verified. The purified hNGF was highly functional as it displayed the ability to induce neuronal differentiation of PC12 cells. Furthermore, it strongly promoted proliferation of TF1 cells, above the levels observed with mouse NGF. Additionally, saliva collected from TG mice and containing unpurified hNGF was able to significantly enhance the growth of TF1 cells. This study not only provides a new and efficient approach for the synthesis of therapeutic hNGF but also supports the concept that salivary gland from TG animals is an efficient system for production of valuable foreign proteins.

Gut microbiota have a complex role on the survivability, digestive physiology, production, and growth performance in animals. Recent studies have emphasized the effects of prebiotics therapy on the gut disease, but the relationship between elephant gut-related diseases and prebiotics remains elusive. Here, a case study was undertaken to evaluate the mechanism of inulin treatment in colic in Asian elephant (Elephas maximus Linnaeus).

Plant-soil feedback (PSF) is conventionally characterized by plant biomass growth, yet it remains unclear how PSF affects plant nutrient acquisition strategies (e.g., nutrient absorption and nutrient resorption) associated with plant growth, particularly under changing soil environments. A greenhouse experiment was performed with seedlings of Pinus elliottii Englem and conditioned soils of monoculture plantations (P. elliottii and Cunninghamia lanceolata Hook). Soil sterilization was designed to test plant phosphorus (P) acquisition strategy with and without native soil fungal communities. Soils from P. elliottii and C. lanceolata plantations were used to explore the specific soil legacy effects on two different P acquisition pathways (absorption and resorption). Phosphorus addition was also applied to examine the separate and combined effects of soil abiotic factors and soil fungal factors on P acquisition pathways. Due to diminished mycorrhizal symbiosis, PSF prompted plants to increasingly rely on P resorption under soil sterilization. In contrast, P absorption was employed preferentially in the heterospecific soil, where species-specific pathogenic fungi could not affect P absorption. Higher soil P availability diluted the effects of soil fungal factors on the trade-off between the two P acquisition pathways in terms of the absolute PSF. Moreover, P addition plays a limited role in terms of the relative PSF and does not affect the direction and strength of relative PSF. Our results reveal the role of PSF in regulating plant P acquisition pathways and highlight the interaction between mycorrhizal and pathogenic fungi as the underlying mechanism of PSF.