Combining ulinastatin with TIENAM improves the outcome of sepsis induced by cecal ligation and puncture in mice by reducing inflammation and regulating immune responses.

Despite the high mortality associated with sepsis, effective and targeted treatments remain scarce. The use of conventional antibiotics such as TIENAM (imipenem and cilastatin sodium for injection, TIE) is challenging because of the increasing bacterial resistance, which diminishes their efficacy and leads to adverse effects. Our previous studies demonstrated that ulinastatin (UTI) exerts a therapeutic impact on sepsis by reducing systemic inflammation and modulating immune responses. In this study, we examined the possibility of administering UTI and TIE after inducing sepsis in a mouse model using cecal ligation and puncture (CLP). We assessed the rates of survival, levels of inflammatory cytokines, the extent of tissue damage, populations of immune cells, microbiota in ascites, and important signaling pathways. The combination of UTI and TIE significantly improved survival rates and reduced inflammation and bacterial load in septic mice, indicating potent antimicrobial properties. Notably, the survival rates of UTI+TIE-treated mice increased from 10 % to 75 % within 168 h compared to those of mice that were subjected to CLP. The dual treatment successfully regulated the levels of inflammatory indicators (interleukin [IL]-6, IL-1ß, and tumor necrosis factor [TNF]-α) and immune cell numbers by reducing B cells, natural killer cells, and TNFR2+ Treg cells and increasing CD8+ T cells. Additionally, the combination of UTI and TIE alleviated tissue damage, reduced bacterial load in the peritoneal cavity, and suppressed the NF-κB signaling pathway. Our findings indicate that UTI and TIE combination therapy can significantly enhance sepsis outcomes by reducing inflammation and boosting the immune system. The results offer a promising therapeutic approach for future sepsis treatment.

Achieving Stable Cycling Performance in a P2-Type Layered Oxide Cathode through a Synergic Li/Zn Doping for Sodium-Ion Batteries.

It has been suggested that sodium layered transition metal oxides could potentially serve as excellent cathodes for sodium-ion batteries (SIBs) because of their appropriate operating potentials and high capacities. However, the growing reliance on energy requirements has necessitated a higher energy density of SIBs. It has been demonstrated that activating oxygen-related activities for SIBs is a viable method to improve energy density. Herein, we suggest applying the synergy of Li and Zn codoping to activate the anionic redox reactions (ARRs) and improve their reversibility in Na0.75Ni0.3Mn0.7O2. The dual ion doping alleviates the phase transition and inhibits the Na+/vacancy arrangements, consequently improving the rate capacity and cyclic stability. The Na0.75Li0.15Ni0.1Zn0.05Mn0.7O2 delivers a discharge capacity of 134 mA h g-1 and no significant capacity loss at 100 mA g-1 between 2 and 4.5 V after 200 cycles. More importantly, the density functional theory (DFT) calculation proves that the codoping strategy triggers more ARRs compared to single-element doping, thereby providing enhanced capacity. The codoping induced oxygen redox strategies will create a new path for rational design of cathodes to enhance the energy density for SIBs.

Cryopreservation of Human Teeth Using Vitrification method with Cryoprotectant Cocktails and N-acetylcysteine for Banking and Clinical Applications.

Preserving freshly-extracted healthy human teeth offers an optional resource for potential tooth transplantation and cell therapy. This study aimed to assess the impact of vitrification, utilizing a blend of cryoprotectant agents and N-acetylcysteine (NAC), on the cryopreservation of periodontal ligament tissues, and investigate the underlying mechanisms of NAC on the tooth cryopreservation. Periodontal ligament cells were isolated from freshly-extracted healthy human permanent teeth, and cell sheets of PDLCs were fabricated. The samples including cell sheets, freshly-extracted human and rat teeth were cryopreserved with or without NAC for three months. The viability, ROS level, gene expressions and microstructure of PDLCs within cell sheets were assessed. The expression of SOD-2, Caspase3, LC3A/B and Catalase were evaluated through western blotting. Histological assessments of cryopreserved cell sheets and teeth were conducted. PDLCs were isolated from cryopreserved teeth, and their immunophenotype and differentiation ability were evaluated. The data was analyzed using one-way analysis of variance. The vitrification method showed good performance in preserving the viability and differentiation potential of PDLCs. Cryopreservation supplemented with NAC improved the survival rate of PDLCs, enhanced osteogenic differentiation ability, upregulated the expression of SOD-2 and Catalase, and inhibited cell apoptosis. Additionally, mRNA sequencing analysis revealed a significant activation of the PI3K-AKT pathway following cryopreservation via vitrification. Adding a PI3K-AKT activator improved the survival rates of PDLCs post-cryopreservation. The vitrification strategy combining various CPAs and NAC proved to be feasible for tooth cryopreservation. Targeting the PI3K-AKT pathway may improve the efficacy of tooth cryopreservation.

Nuclear accumulation of rice UV-B photoreceptors is UV-B- and OsCOP1-independent for UV-B responses.

In plants, the conserved plant-specific photoreceptor UV RESISTANCE LOCUS 8 (UVR8) perceives ultraviolet-B (UV-B) light and mediates UV-B-induced photomorphogenesis and stress acclimation. In this study, we reveal that UV-B light treatment shortens seedlings, increases stem thickness, and enhances UV-B stress tolerance in rice (Oryza sativa) via its two UV-B photoreceptors OsUVR8a and OsUVR8b. Although the rice and Arabidopsis (Arabidopsis thaliana) UVR8 (AtUVR8) photoreceptors all form monomers in response to UV-B light, OsUVR8a, and OsUVR8b function is only partially conserved with respect to AtUVR8 in UV-B-induced photomorphogenesis and stress acclimation. UV-B light and CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1) promote the nuclear accumulation of AtUVR8; by contrast, OsUVR8a and OsUVR8b constitutively localize to the nucleus via their own nuclear localization signals, independently of UV-B light and the RING-finger mutation of OsCOP1. We show that OsCOP1 negatively regulates UV-B responses, and shows weak interaction with OsUVR8s, which is ascribed to the N terminus of OsCOP1, which is conserved in several monocots. Furthermore, transcriptome analysis demonstrates that UV-B-responsive gene expression differs globally between Arabidopsis and rice, illuminating the evolutionary divergence of UV-B light signaling pathways between monocot and dicot plants.

Advances in small-molecule insulin secretagogues for diabetes treatment.

Diabetes, a metabolic disease caused by abnormally high levels of blood glucose, has a high prevalence rate worldwide and causes a series of complications, including coronary heart disease, stroke, peripheral vascular disease, end-stage renal disease, and retinopathy. Small-molecule compounds have been developed as drugs for the treatment of diabetes because of their oral advantages. Insulin secretagogues are a class of small-molecule drugs used to treat diabetes, and include sulfonylureas, non-sulfonylureas, glucagon-like peptide-1 receptor agonists, dipeptidyl peptidase 4 inhibitors, and other novel small-molecule insulin secretagogues. However, many small-molecule compounds cause different side effects, posing huge challenges to drug monotherapy and drug selection. Therefore, the use of different small-molecule drugs must be improved. This article reviews the mechanism, advantages, limitations, and potential risks of small-molecule insulin secretagogues to provide future research directions on small-molecule drugs for the treatment of diabetes.

Metal-free Regioselective Direct C(4)-H Amination of Quinazoline with N-fluorobenzenesulfonimide.

A facile C-H amination of quinazoline employing N-fluorobenzenesulfonimide (NFSI) as the amination source has been disclosed in the absence of any metal, oxidant or additive. The methodology shows a board range of quinazolines with different functional groups in moderate to good yields up to 87%. Furthermore, gram-scale reaction, desulfonylation to amine and synthesis of pharmaceutical intermediate were also investigated, which demonstrates potential applications in medicinal chemistry. A plausible amination mechanism is proposed via F+ transfer accompanied by the removal of one molecule of PhSO2F. DFT studies with experimental work suggest that the mechanism via F+ transfer is more favorable than the free radical one.

Icariin Ameliorates D-galactose-induced Cell Injury in Neuron-like PC12 Cells by Inhibiting MPTP Opening.

OBJECTIVE: Icariin (ICA) has a good neuroprotective effect and can upregulate neuronal basal autophagy in naturally aging rats. Mitochondrial dysfunction is associated with brain aging-related neurodegenerative diseases. Abnormal opening of the mitochondrial permeability transition pore (mPTP) is a crucial factor in mitochondrial dysfunction and is associated with excessive autophagy. This study aimed to explore that ICA protects against neuronal injury by blocking the mPTP opening and down-regulating autophagy levels in a D-galactose (D-gal)-induced cell injury model. METHODS: A cell model of neuronal injury was established in rat pheochromocytoma cells (PC12 cells) treated with 200 mmol/L D-gal for 48 h. In this cell model, PC12 cells were pre-treated with different concentrations of ICA for 24 h. MTT was used to detect cell viability. Senescence associated ß-galactosidase (SA-ß-Gal) staining was used to observe cell senescence. Western blot analysis was performed to detect the expression levels of a senescence-related protein (p21), autophagy markers (LC3B, p62, Atg7, Atg5 and Beclin 1), mitochondrial fission and fusion-related proteins (Drp1, Mfn2 and Opa1), and mitophagy markers (Pink1 and Parkin). The changes of autophagic flow were detected by using mRFP-GFP-LC3 adenovirus. The intracellular ultrastructure was observed by transmission electron microscopy. Immunofluorescence was used to detect mPTP, mitochondrial membrane potential (MMP), mitochondrial reactive oxygen species (mtROS) and ROS levels. ROS and apoptosis levels were detected by flow cytometry. RESULTS: D-gal treatment significantly decreased the viability of PC12 cells, and markedly increased the SA-ß-Gal positive cells as compared to the control group. With the D-gal stimulation, the expression of p21 was significantly up-regulated. Furthermore, D-gal stimulation resulted in an elevated LC3B II/I ratio and decreased p62 expression. Meanwhile, autophagosomes and autolysosomes were significantly increased, indicating abnormal activation of autophagy levels. In addition, in this D-gal-induced model of cell injury, the mPTP was abnormally open, the ROS generation was continuously increased, the MMP was gradually decreased, and the apoptosis was increased. ICA effectively improved mitochondrial dysfunction to protect against D-gal-induced cell injury and apoptosis. It strongly inhibited excessive autophagy by blocking the opening of the mPTP. Cotreatment with ICA and an mPTP inhibitor (cyclosporin A) did not ameliorate mitochondrial dysfunction. However, the protective effects were attenuated by cotreatment with ICA and an mPTP activator (lonidamine). CONCLUSION: ICA inhibits the activation of excessive autophagy and thus improves mitochondrial dysfunction by blocking the mPTP opening.

Biosynthesis of silver nanoparticles using Burkholderia contaminans ZCC and mechanistic analysis at the proteome level.

The biogenic synthesis of silver nanoparticles (AgNPs) by microorganisms has been a subject of increasing attention. Despite extensive studies on this biosynthetic pathway, the mechanisms underlying the involvement of proteins and enzymes in AgNPs production have not been fully explored. Herein, we reported that Burkholderia contaminans ZCC was able to reduce Ag+ to AgNPs with a diameter of (10±5) nm inside the cell. Exposure of B. contaminans ZCC to Ag+ ions led to significant changes in the functional groups of cellular proteins, with approximately 5.72% of the (C-OH) bonds being converted to (C-C/C-H) (3.61%) and CO (2.11%) bonds, and 4.52% of the CO (carbonyl) bonds being converted to (C-OH) bonds. Furthermore, the presence of Ag+ and AgNPs induced the ability of extracellular electron transfer for ZCC cells via specific membrane proteins, but this did not occur in the absence of Ag+ ions. Proteomic analysis of the proteins and enzymes involved in heavy metal efflux systems, protein secretion system, oxidative phosphorylation, intracellular electron transfer chain, and glutathione metabolism suggests that glutathione S-transferase and ubiquinol-cytochrome c reductase iron-sulfur subunit play importance roles in the biosynthesis of AgNPs. These findings contribute to a deeper understanding of the functions exerted by glutathione S-transferase and ferredoxin-thioredoxin reductase iron-sulfur subunits in the biogenesis of AgNPs, thereby hold immense potential for optimizing biotechnological techniques aimed at enhancing the yield and purity of biosynthetic AgNPs.

Research progress on the protective effect of hormones and hormone drugs in myocardial ischemia-reperfusion injury.

Ischemic heart disease (IHD) is a condition where the heart muscle does not receive enough blood flow, leading to cardiac dysfunction. Restoring blood flow to the coronary artery is an effective clinical therapy for myocardial ischemia. This strategy helps lower the size of the myocardial infarction and improves the prognosis of patients. Nevertheless, if the disrupted blood flow to the heart muscle is restored within a specific timeframe, it leads to more severe harm to the previously deprived heart tissue. This condition is referred to as myocardial ischemia/reperfusion injury (MIRI). Until now, there is a dearth of efficacious strategies to prevent and manage MIRI. Hormones are specialized substances that are produced directly into the circulation by endocrine organs or tissues in humans and animals, and they have particular effects on the body. Hormonal medications utilize human or animal hormones as their active components, encompassing sex hormones, adrenaline medications, thyroid hormone medications, and others. While several studies have examined the preventive properties of different endocrine hormones, such as estrogen and hormone analogs, on myocardial injury caused by ischemia-reperfusion, there are other hormone analogs whose mechanisms of action remain unexplained and whose safety cannot be assured. The current study is on hormones and hormone medications, elucidating the mechanism of hormone pharmaceuticals and emphasizing the cardioprotective effects of different endocrine hormones. It aims to provide guidance for the therapeutic use of drugs and offer direction for the examination of MIRI in clinical therapy.

R2* mapping and reduced field-of-view diffusion-weighted imaging for preoperative assessment of nonenlarged lymph node metastasis in rectal cancer.

The aim of the current study is to investigate the diagnostic value of R2* mapping versus reduced field-of-view diffusion-weighted imaging (rDWI) of the primary lesion of rectal cancer for preoperative prediction of nonenlarged lymph node metastasis (NLNM). Eighty-one patients with pathologically confirmed rectal cancer underwent preoperative R2* mapping and rDWI sequences before total mesorectal excisions and accompanying regional lymph node dissections. Two radiologists independently performed whole-tumor measurements of R2* and apparent diffusion coefficient (ADC) parameters on primary lesions of rectal cancer. Patients were divided into positive (NLNM+) and negative (NLNM-) groups based on their pathological analysis. The tumor location, maximum diameter of the tumor, and maximum short diameter of the lymph node were assessed. R2* and ADC, pT stage, tumor grade, status of mesorectal fascia, and extramural vascular invasion were also studied for their potential relationships with NLNM using multivariate logistic regression analysis. The NLNM+ group had significantly higher R2* (43.56 ± 8.43 vs. 33.87 ± 9.57, p < 0.001) and lower ADC (1.00 ± 0.13 vs. 1.06 ± 0.22, p = 0.036) than the NLNM- group. R2* and ADC were correlated to lymph node metastasis (r = 0.510, p < 0.001 for R2*; r = -0.235, p = 0.035 for ADC). R2* and ADC showed good and moderate diagnostic abilities in the assessment of NLNM status with corresponding area-under-the-curve values of 0.795 and 0.636. R2* provided a significantly better diagnostic performance compared with ADC for the prediction of NLNM status (z = 1.962, p = 0.0498). The multivariate logistic regression analysis demonstrated that R2* was a compelling factor of lymph node metastasis (odds ratio = 56.485, 95% confidence interval: 5.759-554.013; p = 0.001). R2* mapping had significantly higher diagnostic performance than rDWI from the primary tumor of rectal cancer in the prediction of NLNM status.

Confidential multiple-input multiple-output optical camera communication aided by a two-dimensional pilot.

Optical camera communication (OCC) has attracted increased attention for its inherent security advantage. However, there still exists the risk of eavesdropping on the broadcasting channel of OCC. To achieve confidential communication, we propose the confidentiality-interference dual light-emitting diode (LED) communication (CIDLC) scheme at the transmitter (TX) and elimination of interference (EI) scheme at the receiver (RX). Meanwhile, interference signals refer to the bit shift of confidential signals. Further, we propose the two-dimensional pilot-aided channel estimation (2D-PACE) scheme to enhance the reliability of multiple-input multiple-output (MIMO) OCC. Experiment results validate the effectiveness of our schemes, which guarantee confidentiality while performing well at a 2 m non-line-of-sight (NLOS) distance. Finally, the communication-illumination integration OCC is constructed via the energy equalization coding (EEC) scheme.

Expression and significance of pigment epithelium-derived factor and vascular endothelial growth factor in colorectal adenoma and cancer.

BACKGROUND: The incidence and mortality of colorectal cancer (CRC) are among the highest in the world, and its occurrence and development are closely related to tumor neovascularization. When the balance between pigment epithelium-derived factors (PEDF) that inhibit angiogenesis and vascular endothelial growth factors (VEGF) that stimulate angiogenesis is broken, angiogenesis is out of control, resulting in tumor development. Therefore, it is very necessary to find more therapeutic targets for CRC for early intervention and later treatment. AIM: To investigate the expression and significance of PEDF, VEGF, and CD31-stained microvessel density values (CD31-MVD) in normal colorectal mucosa, adenoma, and CRC. METHODS: In this case-control study, we collected archived wax blocks of specimens from the Digestive Endoscopy Center and the General Surgery Department of Chengdu Second People's Hospital from April 2022 to October 2022. Fifty cases of specimen wax blocks were selected as normal intestinal mucosa confirmed by electronic colonoscopy and concurrent biopsy (normal control group), 50 cases of specimen wax blocks were selected as colorectal adenoma confirmed by electronic colonoscopy and pathological biopsy (adenoma group), and 50 cases of specimen wax blocks were selected as CRC confirmed by postoperative pathological biopsy after inpatient operation of general surgery (CRC group). An immunohistochemical staining experiment was carried out to detect PEDF and VEGF expression in three groups of specimens, analyze their differences, study the relationship between the two and clinicopathological factors in CRC group, record CD31-MVD in the three groups, and analyze the correlation of PEDF, VEGF, and CD31-MVD in the colorectal adenoma group and the CRC group. The F test or adjusted F test is used to analyze measurement data statistically. Kruskal-Wallis rank sum test was used between groups for ranked data. The chi-square test, adjusted chi-square test, or Fisher's exact test were used to compare the rates between groups. All differences between groups were compared using the Bonferroni method for multiple comparisons. Spearman correlation analysis was used to test the correlation of the data. The test level (α) was 0.05, and a two-sided P< 0.05 was considered statistically significant. RESULTS: The positive expression rate and expression intensity of PEDF were gradually decreased in the normal control group, adenoma group, and CRC group (100% vs 78% vs 50%, χ2 = 34.430, P < 0.001; ++~++ vs +~++ vs -~+, H = 94.059, P < 0.001), while VEGF increased gradually (0% vs 68% vs 96%, χ2 = 98.35, P < 0.001; - vs -~+ vs ++~+++, H = 107.734, P < 0.001). In the CRC group, the positive expression rate of PEDF decreased with the increase of differentiation degree, invasion depth, lymph node metastasis, distant metastasis, and TNM stage (χ2 = 20.513, 4.160, 5.128, 6.349, 5.128, P < 0.05); the high expression rate of VEGF was the opposite (χ2 = 10.317, 13.134, 17.643, 21.844, 17.643, P < 0.05). In the colorectal adenoma group, the expression intensity of PEDF correlated negatively with CD31-MVD (r = -0.601, P < 0.001), whereas VEGF was not significantly different (r = 0.258, P = 0.07). In the CRC group, the expression intensity of PEDF correlated negatively with the expression intensity of CD31-MVD and VEGF (r = -0.297, P < 0.05; r = -0.548, P < 0.05), while VEGF expression intensity was positively related to CD31-MVD (r = 0.421, P = 0.002). CONCLUSION: It is possible that PEDF can be used as a new treatment and prevention target for CRC by upregulating the expression of PEDF while inhibiting the expression of VEGF.

Wet Etching of Silicon in Planar Nanochannels.

Silicon (Si) alkaline etching constitutes a fundamental process in the semiconductor industry. Although its etching kinetics on plain substrates have been thoroughly investigated, the kinetics of Si wet etching in nanoconfinements have yet to be fully explored despite its practical importance in three-dimensional (3-D) semiconductor manufacturing. Herein, we report the systematic study of potassium hydroxide (KOH) wet etching kinetics of amorphous silicon (a-Si)-filled two-dimensional (2-D) planar nanochannels. Our findings reveal that the etching rate would increase with the increase in nanochannel height before reaching a plateau, indicating a strong nonlinear confinement effect. Through investigation using etching solutions with different ionic strengths and/or different temperatures, we further find that both electrostatic interactions and the hydration layer inside the nanoconfinement contribute to the confinement-dependent etching kinetics. Our results offer fresh perspectives into the kinetic study of reactions in nanoconfinements and will shed light on the optimization of etching processes in the semiconductor industry.

AMPA receptor inhibition alleviates inflammatory response and myocardial apoptosis after myocardial infarction by inhibiting TLR4/NF-κB signaling pathway.

Myocardial infarction leads to myocardial inflammation and apoptosis, which are crucial factors leading to heart failure and cardiovascular dysfunction, eventually resulting in death. While the inhibition of AMPA receptors mitigates inflammation and tissue apoptosis, the effectiveness of this inhibition in the pathophysiological processes of myocardial infarction remains unclear. This study investigated the role of AMPA receptor inhibition in myocardial infarction and elucidated the underlying mechanisms. This study established a myocardial infarction model by ligating the left anterior descending branch of the coronary artery in Sprague-Dawley rats. The findings suggested that injecting the AMPA receptor antagonist NBQX into myocardial infarction rats effectively alleviated cardiac inflammation, myocardial necrosis, and apoptosis and improved their cardiac contractile function. Conversely, injecting the AMPA receptor agonist CX546 into infarcted rats exacerbated the symptoms and tissue damage, as reflected by histopathology. This agonist also stimulated the TLR4/NF-κB pathway, further deteriorating cardiac function. Furthermore, the investigations revealed that AMPA receptor inhibition hindered the nuclear translocation of P65, blocking its downstream signaling pathway and attenuating tissue inflammation. In summary, this study affirmed the potential of AMPA receptor inhibition in countering inflammation and tissue apoptosis after myocardial infarction, making it a promising therapeutic target for mitigating myocardial infarction.

Metabolic disorders in prediabetes: From mechanisms to therapeutic management.

Diabetes, one of the world's top ten diseases, is known for its high mortality and complication rates and low cure rate. Prediabetes precedes the onset of diabetes, during which effective treatment can reduce diabetes risk. Prediabetes risk factors include high-calorie and high-fat diets, sedentary lifestyles, and stress. Consequences may include considerable damage to vital organs, including the retina, liver, and kidneys. Interventions for treating prediabetes include a healthy lifestyle diet and pharmacological treatments. However, while these options are effective in the short term, they may fail due to the difficulty of long-term implementation. Medications may also be used to treat prediabetes. This review examines prediabetic treatments, particularly metformin, glucagon-like peptide-1 receptor agonists, sodium glucose cotransporter 2 inhibitors, vitamin D, and herbal medicines. Given the remarkable impact of prediabetes on the progression of diabetes mellitus, it is crucial to intervene promptly and effectively to regulate prediabetes. However, the current body of research on prediabetes is limited, and there is considerable confusion surrounding clinically relevant medications. This paper aims to provide a comprehensive summary of the pathogenesis of pre-diabetes mellitus and its associated therapeutic drugs. The ultimate goal is to facilitate the clinical utilization of medications and achieve efficient and timely control of diabetes mellitus.

Characterization of cells in blood evoked from periapical tissues in immature teeth with pulp necrosis and their potential for autologous cell therapy in Regenerative Endodontics.

OBJECTIVE: The objectives of this study were to isolate, characterize progenitor cells from blood in the root canals of necrotic immature permanent teeth evoked from periapical tissues and evaluate the applicable potential of these isolated cells in Regenerative Endodontics. DESIGN: Ten necrotic immature permanent teeth from seven patients were included. Evoked bleeding from periapical tissues was induced after chemical instrumentation of the root canals. Cells were isolated from the canal blood and evaluated for cell surface marker expression, multilineage differentiation potential, proliferation ability, and target protein expression. Cell sheets formed from these cells were transferred into human root segments, and then transplanted into nude mice. Histological examination was performed after eight weeks. Data analysis was conducted using one-way ANOVA followed by Tukey's post-hoc comparison, considering p < 0.05 as statistically significant. RESULTS: The isolated cells exhibited characteristics typical of fibroblastic cells with colony-forming efficiency, and displayed Ki67 positivity and robust proliferation. Flow cytometry data demonstrated that at passage 3, these cells were positive for CD73, CD90, CD105, CD146, and negative for CD34 and CD45. Vimentin expression indicated a mesenchymal origin. Under differentiation media specific differentiation media, the cells demonstrated osteogenic, adipogenic, and chondrogenic differentiation potential. Subcutaneous root canals with cell sheets of isolated cells in nude mice showed the formation of pulp-like tissues. CONCLUSIONS: This study confirmed the presence of progenitor cells in root canals following evoked bleeding from periapical tissues of necrotic immature teeth. Isolated cells exhibited similar immunophenotype and regenerative potential with dental mesenchymal stromal cells in regenerative endodontic therapy.

Coordinated optimization of the polymerization and transportation processes to enhance the yield of exopolysaccharide heparosan.

Heparosan as the precursor for heparin biosynthesis has attracted intensive attention while Escherichia coli Nissle 1917 (EcN) has been applied as a chassis for heparosan biosynthesis. Here, after uncovering the pivotal role of KfiB in heparosan biosynthesis, we further demonstrate KfiB is involved in facilitating KpsT to translocate the nascent heparosan polysaccharide chain. As a result, an artificial expression cassette KfiACB was constructed with optimized RBS elements, resulting in 0.77 g/L heparosan in shake flask culture. Moreover, in view of the intracellular accumulation of heparosan, we further investigated the effects of overexpression of the ABC transport system proteins on heparosan biosynthesis. By co-overexpressing KfiACB with KpsTME, the heparosan production in flask cultures was increased to 1.03 g/L with an extracellular concentration of 0.96 g/L. Eventually, the engineered strain EcN/pET-kfiACB3-galU-kfiD-glmM/pCDF-kpsTME produced 12.2 g/L heparosan in 5-L fed-batch cultures while the extracellular heparosan was about 11.2 g/L. The results demonstrate the high-efficiency of the strategy for co-optimizing the polymerization and transportation for heparosan biosynthesis. Moreover, this strategy should be also available for enhancing the production of other polysaccharides.

Construction of 5-Aminolevulinic Acid Microbial Cell Factories through Identification of Novel Synthases and Metabolic Pathway Screens and Transporters.

5-Aminolevulinic acid (5-ALA) plays a pivotal role in the biosynthesis of heme and chlorophyll and has garnered great attention for its agricultural applications. This study explores the multifaceted construction of 5-ALA microbial cell factories. Evolutionary analysis-guided screening identified a novel 5-ALA synthase from Sphingobium amiense as the best synthase. An sRNA library facilitated global gene screening that demonstrated that trpC and ilvA repression enhanced 5-ALA production by 74.3% and 102%, respectively. Subsequently, efflux of 5-ALA by the transporter Gdx increased 5-ALA biosynthesis by 25.7%. To mitigate oxidative toxicity, DNA-binding proteins from starved cells were employed, enhancing cell density and 5-ALA titer by 21.1 and 4.1%, respectively. Combining these strategies resulted in an Escherichia coli strain that produced 5-ALA to 1.51 g·L-1 in shake flask experiments and 6.19 g·L-1 through fed-batch fermentation. This study broadens the repertoire of available 5-ALA synthases and transporters and provides a new platform for optimizing 5-ALA bioproduction.

Chronic nicotine exposure elicits pain hypersensitivity through activation of dopaminergic projections to anterior cingulate cortex.

BACKGROUND: Cigarette smoking is commonly reported among chronic pain patients in the clinic. Although chronic nicotine exposure is directly linked to nociceptive hypersensitivity in rodents, underlying neurobiological mechanisms remain unknown. METHODS: Multi-tetrode recordings in freely moving mice were used to test the activity of dopaminergic projections from the ventral tegmental area (VTA) to pyramidal neurones in the anterior cingulate cortex (ACC) in chronic nicotine-treated mice. The VTA→ACC dopaminergic pathway was inhibited by optogenetic manipulation to detect chronic nicotine-induced allodynia (pain attributable to a stimulus that does not normally provoke pain) assessed by von Frey monofilaments (force units in g). RESULTS: Allodynia developed concurrently with chronic (28-day) nicotine exposure in mice (0.36 g [0.0141] vs 0.05 g [0.0018], P<0.0001). Chronic nicotine activated dopaminergic projections from the VTA to pyramidal neurones in the ACC, and optogenetic inhibition of VTA dopaminergic terminals in the ACC alleviated chronic nicotine-induced allodynia in mice (0.06 g [0.0064] vs 0.28 g [0.0428], P<0.0001). Moreover, optogenetic inhibition of Drd2 dopamine receptor signalling in the ACC attenuated nicotine-induced allodynia (0.07 g [0.0082] vs 0.27 g [0.0211], P<0.0001). CONCLUSIONS: These findings implicate a role of Drd2-mediated dopaminergic VTA→ACC pathway signalling in chronic nicotine-elicited allodynia.