High-fat and high-sucrose diet impairs female reproduction by altering ovarian transcriptomic and metabolic signatures.

BACKGROUND: Excessive energy intake in modern society has led to an epidemic surge in metabolic diseases, such as obesity and type 2 diabetes, posing profound threats to women's reproductive health. However, the precise impact and underlying pathogenesis of energy excess on female reproduction remain unclear. METHODS: We established an obese and hyperglycemic female mouse model induced by a high-fat and high-sucrose (HFHS) diet, then reproductive phenotypes of these mice were evaluated by examing sexual hormones, estrous cycles, and ovarian morphologies. Transcriptomic and precise metabolomic analyses of the ovaries were performed to compare the molecular and metabolic changes in HFHS mice. Finally, orthogonal partial least squares discriminant analysis was performed to compare the similarities of traits between HFHS mice and women with polycystic ovary syndrome (PCOS). RESULTS: The HFHS mice displayed marked reproductive dysfunctions, including elevated serum testosterone and luteinizing hormone levels, irregular estrous cycles, and impaired folliculogenesis, mimicking the clinical manifestations of women with PCOS. Precise metabolomic overview suggested that HFHS diet disrupted amino acid metabolism in the ovaries of female mice. Additionally, transcriptional profiling revealed pronounced disturbances in ovarian steroid hormone biosynthesis and glucolipid metabolism in HFHS mice. Further multi-omics analyses unveiled prominent aberration in ovarian arginine biosynthesis pathway. Notably, comparisons between HFHS mice and a cohort of PCOS patients identified analogous reproductive and metabolic signatures. CONCLUSIONS: Our results provide direct in vivo evidence for the detrimental effects of overnutrition on female reproduction and offer insights into the metabolic underpinnings of PCOS.

Abnormalities of retinal function in type 2 diabetes mellitus patients without clinical diabetic retinopathy detected by multifocal electroretinogram.

OBJECTIVE: To study the changes of retinal function in type 2 diabetes mellitus(DM) patients without apparently diabetic retinopathy via multifocal electroretinogram. METHODS: Thirty-six type 2 DM patients (72 eyes) without visible diabetic retinopathy were selected as the experimental group, and thirty-five healthy subjects (70 eyes) were selected as the control group. All subjects were underwent multifocal electroretinogram (mf- ERG). RESULTS: Compared with the control group, the implicit time delay of the P1 wave in the first ring, third ring, fourth ring, and fifth ring of the experimental group was significant (t = -3.154, p = 0.004, t = -8.21, p = 0.000, t = -3.067, p = 0.004, t = -4.443, p = 0.000, respectively). The implicit time of the N1 wave in the fourth- and fifth-ring were also significantly delayed compared with the control group (t = -3.549, p = 0.001, t = 2.961, p = 0.005, respectively). Compared with the control group, the implicit time of the P1 wave and N1 wave in the temporal region of the experimental group were delayed (t = -2.148, p = 0.037, t = -2.834, p = 0.007, respectively). There were no significant difference between the experimental group and the control group of the temporal area in the amplitude density of P1 wave, N1 wave. There was no difference in the implicit time and amplitude density of the N1 and P1 waves in the nasal region between the experimental group and the control group. The multifocal electroretinogram complex parameters showed better specificity and sensitivity in the diagnosis of diabetic retinopathy. CONCLUSION: The multifocal electroretinogram can detect abnormal changes in the retina of type 2 DM patients without visible diabetic retinopathy. The multifocal electroretinogram complex parameter is a potential indicator for the early diagnosis of diabetic retinopathy.

Identification and characterization of two novel ISCR1-associated genes dfrA42 and dfrA43 encoding trimethoprim resistant dihydrofolate reductases.

Two novel ISCR1-associated dfr genes, dfrA42 and dfrA43, were identified from trimethoprim (TMP)-resistant Proteus strains and were shown to confer high level TMP resistance (MIC ≥ 1024 mg/L) when cloned into Escherichia coli These genes were hosted by complex class 1 integrons suggesting their potentials for dissemination. Analysis of enzymatic parameters and TMP affinity were performed, suggesting that the mechanism of TMP resistance for these novel DHFRs is the reduction of binding with TMP.

Pristane cadmium chloride nanoemulsion accelerates the onset of systemic lupus erythematosus in a C57BL/6 mouse model.

OBJECTIVE: To accelerate the onset of systemic lupus erythematosus in C57BL/6 mice by injecting cadmium chloride nanoemulsion and shorten the traditional modeling time. METHODS: Pristane cadmium chloride nanoemulsion was prepared, and 66 C57BL/6 mice were randomly divided into four groups. The pristane group was intraperitoneally injected with 0.6 mL of pristane blank nanoemulsion, the model group was injected with 0.6 mL of pristane cadmium chloride nanoemulsion, the Cadmium chloride control group was injected with 0.6 mL of cadmium chloride nanoemulsion, and the control group was injected with the same amount of 0.9% sodium chloride solution. Urine protein content, anti-dsDNA antibody content, Th1 cell/Th2 cell ratio, and kidney staining were detected in each group. RESULTS: The model group began to develop disease in the 4th week, the anti-dsDNA antibody level reached 566.71 ± 1.44 ng/L, and the proteinuria reached 245.38 ± 30.54 ng/mL. The model group showed an onset at least 5 weeks earlier than that in the pristane group. There was no significant difference in anti-dsDNA antibody content between Cadmium chloride control group and blank group. At the 12th week, the Th1/Th2 cell ratio in the model group significantly decreased, and the pathological changes in the kidneys were consistent with the typical manifestations of lupus in mouse models. CONCLUSION: These results suggest that cadmium chloride promotes earlier onset of pristane-induced systemic lupus erythematosus in a C57BL/6 mouse model.

Co-occurring tree species drive arbuscular mycorrhizal fungi diversity in tropical forest.

It is still uncertain whether environment or host plant species is more important in determining AMF diversity; although, plant roots are usually associated with abundant AMF species in different environments. This study explored the effect of plant species and environmental factors on AMF diversity based on three co-occurring tree species (Glochidion coccineum, Schefflera octophylla, and Schima superba) on six elevations of Mt. Jianfengling. A total of 185 OTUs (operational taxonomic units) of AMF were found in the three co-occurring dominant tree species. Of which 109 unique OTUs were identified in the three co-occurring plant species, which accounted for the total number of 58.92%. Forty-five OTUs were shared by the three co-occurring tree species, accounting for a total number of 24.32%. The plant species of Schefflera octophylla was identified as having the highest AMF diversity with the largest number of OTUs of 143. The fungi in the genus of Glomus were the dominant AMF species in the three co-occurring tree species. AMF communities and diversity are quite different, either within different plant species at the same elevation or within the same plant species at different elevations. However, the altitude had no significant effect on the ACE index. Therefore, the results suggest that plant species have a more important effect on AMF diversity and community composition.

[Design and implementation of a modular pulse wave preprocessing and analysis system based on a new detection algorithm].

As one of the standard electrophysiological signals in the human body, the photoplethysmography contains detailed information about the blood microcirculation and has been commonly used in various medical scenarios, where the accurate detection of the pulse waveform and quantification of its morphological characteristics are essential steps. In this paper, a modular pulse wave preprocessing and analysis system is developed based on the principles of design patterns. The system designs each part of the preprocessing and analysis process as independent functional modules to be compatible and reusable. In addition, the detection process of the pulse waveform is improved, and a new waveform detection algorithm composed of screening-checking-deciding is proposed. It is verified that the algorithm has a practical design for each module, high accuracy of waveform recognition and high anti-interference capability. The modular pulse wave preprocessing and analysis software system developed in this paper can meet the individual preprocessing requirements for various pulse wave application studies under different platforms. The proposed novel algorithm with high accuracy also provides a new idea for the pulse wave analysis process.

LS-106, a novel EGFR inhibitor targeting C797S, exhibits antitumor activities both in vitro and in vivo.

With the wide clinical use of the third-generation epidermal growth factor receptor (EGFR) inhibitor osimertinib for the treatment of EGFR-mutated non-small cell lung cancer (NSCLC), acquired resistance caused by EGFR C797S tertiary mutation has become a concern. Therefore, fourth-generation EGFR inhibitors that could overcome this mutation have gained increasing attention in recent years. Here, we identified LS-106 as a novel EGFR inhibitor against C797S mutation and evaluated its antitumor activity both in vitro and in vivo. In cell-free assay, LS-106 potently inhibited the kinase activities of EGFR19del/T790M/C797S and EGFRL858R/T790M/C797S with IC50 values of 2.4 nmol/L and 3.1 nmol/L, respectively, which was more potent than osimertinib. Meanwhile, LS-106 exhibited comparable kinase inhibitory effect to osimertinib on EGFRL858R/T790M and wild-type EGFR. Results from cellular experiments demonstrated that LS-106 potently blocked the phosphorylation of EGFR C797S triple mutations in the constructed BaF3 cells that highly expressed EGFR19del/T790M/C797S or EGFRL858R/T790M/C797S , and thus inhibited the proliferation of these cells. We also constructed tumor cells harboring EGFR19del/T790M/C797S (named PC-9-OR cells) using the CRISPR/Cas9 system and found that LS-106 markedly suppressed the activation of EGFR19del/T790M/C797S and the proliferation of PC-9-OR cells. Moreover, cells harboring EGFR19del/T790M/C797S underwent remarkable apoptosis upon LS-106 treatment. In vivo experiments further demonstrated that oral administration of LS-106 caused significant tumor regression in a PC-9-OR xenograft model, with a tumor growth inhibition rate (TGI) of 83.5% and 136.6% at doses of 30 and 60 mg/kg, respectively. Taken together, we identified LS-106 as a novel fourth-generation EGFR inhibitor against C797S mutation and confirmed its preclinical antitumor effects in C797S-triple-mutant tumor models.

TfOH-Promoted Decyanative Cyclization toward the Synthesis of 2,1-Benzisoxazoles.

A novel solvent-free, TfOH-promoted decyanative cyclization approach for the synthesis of 2,1-benzisoxazoles has been developed. The reactions are complete instantly at room temperature and result in the formation of the desired 2,1-benzisoxazoles in a 34-97% isolated yield.

Synthesis of Disulfide Surrogate Peptides Incorporating Large-Span Surrogate Bridges Through a Native-Chemical-Ligation-Assisted Diaminodiacid Strategy.

The use of synthetic bridges as surrogates for disulfide bonds has emerged as a practical strategy to obviate the poor stability of some disulfide-containing peptides. However, peptides incorporating large-span synthetic bridges are still beyond the reach of existing methods. Herein, we report a native chemical ligation (NCL)-assisted diaminodiacid (DADA) strategy that enables the robust generation of disulfide surrogate peptides incorporating surrogate bridges up to 50 amino acids in length. This strategy provides access to some highly desirable but otherwise impossible-to-obtain disulfide surrogates of bioactive peptide. The bioactivities and structures of the synthetic disulfide surrogates were verified by voltage clamp assays, NMR, and X-ray crystallography; and stability studies established that the disulfide replacements effectively overcame the problems of disulfide reduction and scrambling that often plague these pharmacologically important peptides.

The diversity of glycosylation of cellobiohydrolase I from Trichoderma reesei determined with mass spectrometry.

Cellulases are glycosylated enzymes that have wide applications in fields like biofuels. It has been widely accepted that glycosylation of cellulases impact their performance. Trichoderma reesei is the most important cellulase-producer and cellobiohydrolase I (CBHI) is the most important cellulase from T. reesei. Therefore, the glycosylation of T. reesei CBHI has been a focus of research. However, investigations have been focused on N-glycosylation of three of the four potential glycosylation sites, as well as O-glycosylation on the linker region, while a full picture of glycosylation of T. reesei CBHI is still needed. In this work, with extensive mass spectrometric investigations on CBHI from two T. reesei strains grown under three conditions, several new discoveries were made: 1) N45 and N64 are N-glycosylated with high mannose type glycans; 2) the catalytic domain of CBHI is extensively O-glycosylated with hexoses and N-acetylhexosamines; 3) experimental evidence on the mannosylation of carbohydrate binding domain (other than the linker adjacent region) was found. With structural analysis, we found several glycosylation sites (such as T383, S8, and S46) are located at the openings of the substrate-binding tunnel, and potentially involve in the binding of cellulose. These investigations provide a full and comprehensive picture on the glycosylation of CBHI from T. reesei, which benefits the engineering of CBHI by raising potential sites for modification.

Prediction of virus-host infectious association by supervised learning methods.

BACKGROUND: The study of virus-host infectious association is important for understanding the functions and dynamics of microbial communities. Both cellular and fractionated viral metagenomic data generate a large number of viral contigs with missing host information. Although relative simple methods based on the similarity between the word frequency vectors of viruses and bacterial hosts have been developed to study virus-host associations, the problem is significantly understudied. We hypothesize that machine learning methods based on word frequencies can be efficiently used to study virus-host infectious associations. METHODS: We investigate four different representations of word frequencies of viral sequences including the relative word frequency and three normalized word frequencies by subtracting the number of expected from the observed word counts. We also study five machine learning methods including logistic regression, support vector machine, random forest, Gaussian naive Bayes and Bernoulli naive Bayes for separating infectious from non-infectious viruses for nine bacterial host genera with at least 45 infecting viruses. Area under the receiver operating characteristic curve (AUC) is used to compare the performance of different machine learning method and feature combinations. We then evaluate the performance of the best method for the identification of the hosts of contigs in metagenomic studies. We also develop a maximum likelihood method to estimate the fraction of true infectious viruses for a given host in viral tagging experiments. RESULTS: Based on nine bacterial host genera with at least 45 infectious viruses, we show that random forest together with the relative word frequency vector performs the best in identifying viruses infecting particular hosts. For all the nine host genera, the AUC is over 0.85 and for five of them, the AUC is higher than 0.98 when the word size is 6 indicating the high accuracy of using machine learning approaches for identifying viruses infecting particular hosts. We also show that our method can predict the hosts of viral contigs of length at least 1kbps in metagenomic studies with high accuracy. The random forest together with word frequency vector outperforms current available methods based on Manhattan and [Formula: see text] dissimilarity measures. Based on word frequencies, we estimate that about 95% of the identified T4-like viruses in viral tagging experiment infect Synechococcus, while only about 29% of the identified non-T4-like viruses and 30% of the contigs in the study potentially infect Synechococcus. CONCLUSIONS: The random forest machine learning method together with the relative word frequencies as features of viruses can be used to predict viruses and viral contigs for specific bacterial hosts. The maximum likelihood approach can be used to estimate the fraction of true infectious associated viruses in viral tagging experiments.

Effect of exercise on postoperative recovery of patients with non-small cell lung cancer: a systematic review and meta-analysis.

Patients with non-small cell lung cancer (NSCLC) in the postoperative recovery period often experience reduced exercise capacity and impaired lung function, which affects their overall quality of life. This paper investigated the effect of exercise interventions on exercise capacity, lung function, quality of life, and symptoms in these patients. METHODS: We performed a literature search across Cochrane, Embase, PubMed, Web of Science, and EBSCO databases were comprehensively searched for randomized controlled trials (RCTs) from inception to September 2023, all English RCTs were eligible if they assessed the effects of exercise interventions on postoperative NSCLC patients. RESULTS: Twelve articles met our inclusion criteria, evidencing that exercise interventions could significantly improve the functional capacity of NSCLC patients in postoperative recovery. Notably, Forced Expiratory Volume in 1 s (FEV1) was improved, indicating enhanced lung function. Furthermore, exercise improved the physical and mental health scores of SF-36, along with increased quadriceps strength and relieved dyspnea. However, fatigue levels were not significantly changed. CONCLUSIONS: Exercise interventions of NSCLC patients in the postoperative recovery are associated with improved functional capacity, lung function, quality of life, and quadriceps strength, as well as alleviated symptoms of dyspnea. These findings underscore the potential benefits of incorporating exercise into postoperative care for NSCLC patients. Nonetheless, further large-scale RCTs are required to solidify the evidence base on the clinical outcomes of exercise following pneumonectomy.

Metal-organic-framework-confined quantum dots enhance photocurrent signals: A molecularly imprinted photoelectrochemical cathodic sensor for rapid and sensitive tetracycline detection.

BACKGROUND: Tetracycline (TC), a cost-effective broad-spectrum antibacterial drug, has been excessively utilized in the livestock and poultry industry, leading to a serious overabundance of TC in livestock wastewater. However, conventional analytical methods such as liquid chromatography and gas chromatography face challenges in achieving sensitive detection of trace amounts of TC in complex substrates. Therefore, it is imperative to develop a highly sensitive and anti-interference analytical method for the detection of tetracycline in livestock wastewater. RESULTS: A porphyrin-based MOF (PCN-224)-confined carbon dots (CDs) material (CDs@PCN-224) was synthesized by a "bottle-around-ship" strategy. The reduced carrier migration distance is conducive to the separation of electron-hole pairs and enhanced the photocurrent signal due to the tight coupling of CDs and PCN-224. Further, molecularly imprinted polymer (MIP) was synthesized by rapid in-situ UV-polymerization and employed as a recognition element. The specific recognition of the target by imprinted cavities blocks electron transfer, resulting in a "turn off" response signal, thus realizing the selective detection of TC. Under optimal conditions, the constructed MIP-PEC cathodic sensor detected 1.00 × 10-12 M to 1.00 × 10-7 M of TC sensitively, with a limit of detection of 3.72 × 10-13 M. In addition, the proposed MIP-PEC sensor demonstrated good TC detection performance in actual livestock wastewater. SIGNIFICANCE: The strategy based on MOF pore-confined quantum dots can effectively enhance the photocurrent response of the photosensitive substrate. Simultaneously, the MIP constructed by in-situ rapid UV-polymerization showed excellent anti-interference and reusable properties. This work provides a promising MIP-PEC cathodic sensing method for the rapid and sensitive detection of antibiotics in complex-matrix environmental samples.

Geniposide ameliorates psoriatic skin inflammation by inhibiting the TLR4/MyD88/NF-κB p65 signaling pathway and MMP9.

Psoriasis is an incurable immune-mediated disease affecting the skin or the joints. There are continuing studies on drugs for psoriasis prevention and treatment. This research found that Geniposide (GE) significantly thinned IMQ mice's skin lesions, reduced the scales, and lowered the presence of inflammatory cells in the pathology in a dose-dependent manner. GE inhibited IL-23, IL-22, IL-17A, IL-12, IL-6, and TNF-α levels in psoriatic mice serum. AKT1, TNF, TLR4, MMP9, MAPK3, and EGFR were selected as the top 6 targets of GE against psoriasis via network pharmacology, and GE-TLR4 has the most robust docking score value by molecular docking. Taken together, GE significantly inhibited TLR4 and MMP9 protein expression and influenced MyD88/NF-κB p65 signaling pathway. Finally, TLR4 was verified as the critical target of GE, which engaged in immunomodulatory activities and reduced MMP9 production in LPS and TAK-242-induced HaCaT cells. GE had a medium affinity for TLR4, and the KD value was 1.06 × 10-5 M. GE is an effective treatment and preventative strategy for psoriasis since it impacts TLR4.

A Systematic Narrative Review of Implementation, Processes, and Outcomes of Human Library.

First started in Denmark in 2000, Human Library (HL) has been adopted by different communities around the world. It is an innovative approach that engages "readers" from the general public to have collaborative conversations with "books" from minority or marginalized communities to learn about their lived experiences and reduce public stigma and stereotypes. While the HL is popularized, its research base and implementation structure remain limited. This systematic narrative review aims to review the HL literature to (1) summarize the design, implementation, processes, and outcomes of existing HL programs and (2) synthesize recommendations for future implementation of more effective, ethical, and sensible HL. A systematic search in eight electronic databases yielded 23 journal articles and book chapters about HL published from 2010 to 2022. The identified literature demonstrated variations in format, venue, scale, preparation, and recruitment. A wide range of books with different social identities and from different cultural groups were reported, while readers were mostly from university and school communities. Reduced prejudices and improved attitudes were reported in readers, while both readers and books reported various facets of personal growth. Future HL using implementation guidelines with clearly articulate ethical considerations and diverse rigorous research methodologies are recommended.

AMF Inoculation Alleviates Molybdenum Toxicity to Maize by Protecting Leaf Performance.

The use of arbuscular mycorrhizal fungi (AMF) is a vital strategy for enhancing the phytoremediation of heavy metals. However, the role of AMF under molybdenum (Mo) stress is elusive. A pot culture experiment was conducted to explore the effects of AMF (Claroideoglomus etunicatum and Rhizophagus intraradices) inoculation on the uptake and transport of Mo and the physiological growth of maize plants under different levels of Mo addition (0, 100, 1000, and 2000 mg/kg). AMF inoculation significantly increased the biomass of maize plants, and the mycorrhizal dependency reached 222% at the Mo addition level of 1000 mg/kg. Additionally, AMF inoculation could induce different growth allocation strategies in response to Mo stress. Inoculation significantly reduced Mo transport, and the active accumulation of Mo in the roots reached 80% after inoculation at the high Mo concentration of 2000 mg/kg. In addition to enhancing the net photosynthetic and pigment content, inoculation also increased the biomass by enhancing the uptake of nutrients, including P, K, Zn, and Cu, to resist Mo stress. In conclusion, C. etunicatum and R. intraradices were tolerant to the Mo stress and could alleviate the Mo-induced phytotoxicity by regulating the allocation of Mo in plants and improving photosynthetic leaf pigment contents and the uptake of nutrition. Compared with C. etunicatum, R. intraradices showed a stronger tolerance to Mo, which was manifested by a stronger inhibition of Mo transport and a higher uptake of nutrient elements. Accordingly, AMF show potential for the bioremediation of Mo-polluted soil.

Intragastric administration of prednisone acetate induced impairment of hippocampal long-term potentiation.

Prednisone acetate (PA) has many adverse side effects despite the fact that oral administration of PA is widely administrated in the clinic. However, it is unknown whether PA can cause hippocampal long-term potentiation (LTP) impairment. Therefore, in our study, PA (5 mg/kg·d) through intragastric administration (gavage) was applied to establish a model of impaired hippocampal LTP in C57BL/6 mice, and the method was evaluated by comparing with another method to establish LTP impairment through subcutaneous injection of corticosterone (CORT, 50 mg/kg·d). First, our results showed PA caused a more significant decrease in population spike (PS, %) after high-frequency stimulation (HFS) than CORT, demonstrating PA induced impairment of hippocampal LTP more successfully than CORT. Second, PA caused poorer performance of memory than CORT. Third, PA caused more serious lesions and loss of the granule cell in the dentate gyrus than CORT. Finally, PA caused lower levels of glutamic acid (Glu), N-methyl-d-aspartate receptors (NMDARs) and gamma-aminobutyric acid (GABA) than CORT. All in all, PA (5 mg/kg·d) through intragastric administration (gavage) induced LTP impairment in the hippocampus more successfully than CORT. The neuronal lesions in the dentate gyrus and the consequent decrease of Glu and NMDARs (especially NMDAR2A) may be the cause of LTP impairment.

Surfaces of gymnastic equipment as reservoirs of microbial pathogens with potential for transmission of bacterial infection and antimicrobial resistance.

Gymnastic equipment surfaces are shared by many people, and could mediate the transfer of bacterial pathogens. To better understand this detrimental potential, investigations on the reservoirs of bacterial pathogens and antimicrobial resistance on the surfaces of gymnastic equipment were performed by analyzing the bacterial community structures, prevalence of viable bacteria, and presence of antimicrobial resistance on both indoor and outdoor gymnastic facilities. The results of high-throughput 16S rDNA amplicon sequencing showed that Gram-positive bacteria on the surfaces of indoor gymnastic equipment significantly enriched, including the opportunistic pathogen Staphylococcus strains, while Enterobacteriaceae significantly enriched on surfaces of outdoor gymnastic equipment. The analysis of α-diversities showed a higher richness and diversity for bacterial communities on the surfaces of gymnastic equipment than the environment. Analysis of ß-diversities showed that the bacterial communities on the surfaces of gymnastic equipment differ significantly from environmental bacterial communities, while the bacterial communities on indoor and outdoor equipment are also significantly different. Thirty-four bacterial isolates were obtained from the surfaces of gymnastic equipment, including three multidrug Staphylococcus and one multidrug resistant Pantoea. In particular, Staphylococcus hemolyticus 5-6, isolated from the dumbbell surface, is a multidrug resistant, hemolytic, high- risk pathogen. The results of quantitative PCR targeting antibiotic resistance related genes (intI1, sul1 and bla TEM) showed that the abundances of sul1 and bla TEM genes on the surfaces of gymnastic equipment are higher than the environment, while the abundances of sul1 gene on indoor equipment are higher than outdoor equipment. These results lead to the conclusion that the surfaces of gymnastic equipment are potential dissemination pathways for highly dangerous pathogens as well as antimicrobial resistance, and the risks of indoor equipment are higher than outdoor equipment.

Cryptosporidium parvum maintains intracellular survival by activating the host cellular EGFR-PI3K/Akt signaling pathway.

Autophagy is a critical cellular mechanism in helping infected cells remove intracellular pathogens and is countered by pathogens maintaining intracellular survival by regulating autophagy through the manipulation of the host cellular signal transduction pathway. Cryptosporidium parvum is a zoonotic intracellular but extracytoplasmic protozoon that causes diarrhea in infants and young children worldwide. However, it is still unclear how Cryptosporidium adapts to intracellular survival. In the present study, we demonstrated that C. parvum could activate the EGFR-PI3K/Akt signaling pathway to promote intracellular survival in HCT-8 cells. The western blot results showed that C. parvum induced EGFR and Akt phosphorylation in HCT-8 cells. The EGFR inhibitor AG1478 decreased EGFR and Akt phosphorylation, and the PI3K inhibitor LY294002 impaired Akt phosphorylation induced by C. parvum in HCT-8 cells. Inhibition of EGFR or Akt decreased the number of intracellular parasites. Second, low-dose infection of C. parvum triggered complete autophagy and enhanced autophagic flux in HCT-8 cells. The expressions of mTOR and p62 were decreased, and the expressions of LC3 and Beclin1 were increased in C. parvum-infected HCT-8 cells. Transfection with siBeclin1 or siATG7 reduced LC3 accumulation, while lysosome inhibitor E64d+pepA increased LC3 accumulation induced by C. parvum in HCT-8 cells. Intracellular parasite proliferation was decreased when treated with autophagy inducer rapamycin, whereas autophagy inhibitor 3-MA, E64d+pep A, siBeclin1 or siATG7 increased intracellular parasites. Third, C. parvum inhibited autophagy killing to promote its own intracellular survival by activating EGFR-Akt signaling pathway. The EGFR inhibitor AG1478 enhanced autophagic flux, and Akt inhibitor IV increased LC3 accumulation and inhibited C. parvum proliferation in HCT-8 cells. Akt inhibitor IV-inhibited C. parvum proliferation was attenuated by E64d+pepA. In summary, C. parvum could maintain intracellular survival by inhibiting autophagy via EGFR-PI3K/Akt pathway. These results revealed a new mechanism for the interaction of C. parvum with host cells.

Comparative Analysis of Transcriptomic Response of Escherichia coli K-12 MG1655 to Nine Representative Classes of Antibiotics.

The use of antibiotics leads to strong stresses to bacteria, leading to profound impact on cellular physiology. Elucidating how bacteria respond to antibiotic stresses not only helps us to decipher bacteria's strategies to resistant antibiotics but also assists in proposing targets for antibiotic development. In this work, a comprehensive comparative transcriptomic analysis on how Escherichia coli responds to nine representative classes of antibiotics (tetracycline, mitomycin C, imipenem, ceftazidime, kanamycin, ciprofloxacin, polymyxin E, erythromycin, and chloramphenicol) was performed, aimed at determining and comparing the responses of this model organism to antibiotics at the transcriptional level. On average, 39.71% of genes were differentially regulated by antibiotics at concentrations that inhibit 50% growth. Kanamycin leads to the strongest transcriptomic response (76.4% of genes regulated), whereas polymyxin E led to minimal transcriptomic response (4.7% of genes regulated). Further GO, KEGG, and EcoCyc enrichment analysis found significant transcriptomic changes in carbon metabolism, amino acid metabolism, nutrient assimilation, transport, stress response, nucleotide metabolism, protein biosynthesis, cell wall biosynthesis, energy conservation, mobility, and cell-environmental communications. Analysis of coregulated genes led to the finding of significant reduction of sulfur metabolism by all antibiotics, and analysis of transcription factor-coding genes suggested clustered regulatory patterns implying coregulation. In-depth analysis of regulated pathways revealed shared and unique strategies of E. coli resisting antibiotics, leading to the proposal of four different strategies (the pessimistic, the ignorant, the defensive, and the invasive). In conclusion, this work provides a comprehensive analysis of E. coli's transcriptomic response to antibiotics, which paves the road for further physiological investigation. IMPORTANCE Antibiotics are among the most important inventions in the history of humankind. They are the ultimate reason why bacterial infections are no longer the number one threat to people's lives. However, the wide application of antibiotics in the last half a century has led to aggravating antibiotic resistance, weakening the efficacy of antibiotics. To better comprehend the ways bacteria deal with antibiotics that may eventually turn into resistance mechanisms, and to identify good targets for potential antibiotics, knowledge on how bacteria regulate their physiology in response to different classes of antibiotics is needed. This work aimed to fill this knowledge gap by identifying changes of bacterial functions at the transcription level and suggesting strategies of bacteria to resist antibiotics.