Research on the Interaction Mechanism and Structural Changes in Human Serum Albumin with Hispidin Using Spectroscopy and Molecular Docking.

The interaction between human serum albumin (HSA) and hispidin, a polyketide abundantly present in both edible and therapeutic mushrooms, was explored through multispectral methods, hydrophobic probe assays, location competition trials, and molecular docking simulations. The results of fluorescence quenching analysis showed that hispidin quenched the fluorescence of HSA by binding to it via a static mechanism. The binding of hispidin and HSA was validated further by synchronous fluorescence, three-dimensional fluorescence, and UV/vis spectroscopy analysis. The apparent binding constant (Ka) at different temperatures, the binding site number (n), the quenching constants (Ksv), the dimolecular quenching rate constants (Kq), and the thermodynamic parameters (∆G, ∆H, and ∆S) were calculated. Among these parameters, ∆H and ∆S were determined to be 98.75 kJ/mol and 426.29 J/(mol·K), respectively, both exhibiting positive values. This observation suggested a predominant contribution of hydrophobic forces in the interaction between hispidin and HSA. By employing detergents (SDS and urea) and hydrophobic probes (ANS), it became feasible to quantify alterations in Ka and surface hydrophobicity, respectively. These measurements confirmed the pivotal role of hydrophobic forces in steering the interaction between hispidin and HSA. Site competition experiments showed that there was an interaction between hispidin and HSA molecules at site I, which situates the IIA domains of HSA, which was further confirmed by the molecular docking simulation.

The phosphorylation of Tudor-SN mediated by JNK is involved in the regulation of milk protein synthesis induced by prolactin in BMECs.

Tudor staphylococcal nuclease (Tudor-SN) is a multifunctional protein involved in a variety of cellular processes and plays a critical role in the regulation of gene expression. Recently, Tudor-SN was found to be upregulated in mammary epithelial cells during lactation in response to prolactin, which further to regulate milk protein synthesis. However, the detailed regulatory mechanism of Tudor-SN to milk protein still remains to be elucidated. In our study, we observed that the levels of Tudor-SN and phosphor-Tudor-SN (Thr103) were both enhanced upon prolactin stimulation. Immunofluorescence assays demonstrated that prolactin treatment facilitated the nuclear transport of Tudor-SN. Further study revealed that the phosphorylation of Tudor-SN was depended on activated JNK. Coimmunoprecipitation assays disclosed that Tudor-SN might be phosphorylated directly by JNK. Using gene mutation assays, we further discovered that mutation of Thr to Ala at site of 103 prevented the nuclear transport of Tudor-SN. Thus, these results suggested the essential mechanism of the activated Tudor-SN in milk protein regulation in response to prolactin, which may provide some new sights into improve milk protein production.

Fusion expression of cecropin B-like antibacterial peptide in Pichia GS115 and its antibacterial mechanism.

OBJECTIVES: To establish an efficient expression system for a fusion protein of glutathione S-transferase and cecropin B (GST-CB) and to clarify the antibacterial mechanism of CB. RESULTS: The optimal incubation time and methanol concentration for induced expression of CB were 36 h and 1 % w/v, respectively. The yield of GST-CB was 2.2 g/l. The minimum inhibitory concentrations of GST-CB towards Staphylococcus aureus subsp. saprophyticus (ATCC 15305) and Escherichia coli strain CFT073 were 250 and 125 µg/ml, respectively. Notably, mutations of proline 24 (P24) in CB produced a polypeptide without antimicrobial activity. CONCLUSION: The fusion protein GST-CB, which has a broad spectrum antimicrobial activity, can be abundantly expressed in Pichia pastoris GS115, and P24 may be an important amino acid for the antimicrobial activity of GST-CB.

[Antibacterial effect of self-etching adhesive systems on Streptococcus mutans].

OBJECTIVE: To investigate the antibacterial effect of different self-etching adhesive systems against Streptococcus mutans (S. mutans). METHODS: Six reagents Clearfil(TM) SE Bond primer (SP), Clearfil(TM) SE Bond adhesive (SA),Clearfil(TM) Protect Bond primer (PP), which contained antibacterial monomer methacryloyloxydodecylpyridinium bromide (MDPB), ClearfilTM Protect Bond adhesive (PA), positive control chlorhexidine acetate [CHX, 1% (mass fraction)], and negative control phosphate buffer solution (PBS) were selected. They were mixed with S. mutans for 30 s respectively, then colony-forming units (CFU) were counted after incubated for 48 h on brain heart infusion (BHI) agar medium. The 6 reagents were applied to the sterile paper discs, and distributed onto the BHI agar medium with S. mutans and incubated for 24 h, then the inhibition zones were observed. CHX, PBS, PP, and SP were added on the dentin with artificial caries induced by S. mutans and kept for 30 s, then confocal laser scanning microscope (CLSM) was used to observe the live and dead bacteria after staining. The ratio of live to dead bacteria was calculated. PP+PA and SP+SA were applied on the dentin according to the manual and light cured. S. mutans were incubated on the samples for 2 h, ultrasonically treated and incubated on BHI agar medium for 48 h, then CFU was counted. The data were analyzed by non-parametric analysis and one-way ANOVA. RESULTS: Compared with PBS, the PP, SP, PA, SA and CHX showed the antibacterial effect on free S. mutans (P<0.05); SP and PP showed stronger antibacterial effect than PA, SA and CHX (P<0.05). CHX, SP and PP presented inhibition zones, while PBS, SA and PA did not. Compared with PBS, the CHX, SP and PP could lower the ratio of the live to dead bacteria significantly (P<0.05). Cured self-etching adhesive systems did not show any antibacterial effect on the free S. mutans. CONCLUSION: The primer of self-etching adhesives Clearfil(TM) SE Bond and Clearfil(TM) Protect Bond showed significant antibacterial effect on free and attached S. mutans. The adhesive only showed antibacterial effect on free S. mutans before light-cured polymerization. After being cured, the self-etching adhesive systems did not show antibacterial effect anymore.

Nobiletin inhibited hypoxia-induced epithelial-mesenchymal transition of lung cancer cells by inactivating of Notch-1 signaling and switching on miR-200b.

Epithelial-mesenchymal transition (EMT) is an early step in the process of tumor metastasis. It is well known that tumor microenvironment affects malignancy in various carcinomas; in particular, that hypoxia induces EMT. Deregulated notch signaling also contributes a lot to the development of EMT in lung cancer. In this study, we investigated the use of Notch-1-inhibiting compound as novel therapeutic candidates to regulate hypoxia-induced EMT in lung cancer cells. According to previous screening, nobiletin was selected as a Notch-1 inhibitor. Hypoxia-induced EMT was characteristic of increased N-cadherin & vimentin expressions and decreased E-cadherin expressions. Treatment with nobiletin notably attenuated hypoxia-induced EMT, invasion and migration in H1299 cells, accompanied with reduced Notch-1, Jagged1/2 expressions and its downstream genes Hey-1 and Hes-1. Nobiletin treatment also promoted tumorsuppressive miR-200b level. Moreover, notch-1 siRNA prevented hypoxia-mediated cell migration and decreased Twist1, Snail1, and ZEB1/2 expressions, which are key EMT markers. Re-expression of miR-200b blocked hypoxia-induced EMT and cell invasion. Our findings suggest that downregulation of Notch-1 and reexpression of miR-200b by nobiletin might be a novel remedy for the therapy of lung cancer.

[Real temperature of the continuous-wave pluggers].

OBJECTIVE: To measure the real temperatures on the pluggers of three continuous-wave devices, and to provide theoretical reference to evaluate thermal damage and heat's influence on the filling materials. METHODS: The dual channel K type thermocouple was contacted to various sizes' pluggers in three different continuous-wave devices (BeeFill, Elements, B&L), and the highest temperatures at different points (tip, and 2 mm, 5 mm, 10 mm from the tip) of the pluggers (preset temperature was 200 °C) were recorded. The measurements were performed 5 times. T-test was used to compare the real temperatures at the tips with that set on the display and one-way ANOVA was used to compare the temperatures of the pluggers in different devices, sizes and points. RESULTS: The highest temperature was at the tip of BeeFill 40/0.03 plugger (198.7±7.7) °C, but there was on statistical differences between that and the preset temperature 200 °C. The temperatures of the remaining pluggers were obviously lower than 200 °C (P<0.05). The lowest temperature of the pluggers was detected at 10 mm from the tip of BeeFill 60/0.06 plugger (69.9±4.0) °C. The highest temperature of each plugger was detected at the tip or 2 mm from the tip (112.1 to 198.7 °C,and the median was 140.8 °C). CONCLUSION: The real temperature of most continuous-wave pluggers included in this study is below the set temperature 200 °C.

[Synergistic effects of lysozyme with EDTA-2Na on antibacterial activity].

OBJECTIVE: To evaluate the synergistic antibacterial effects of lysozyme with ethylenediaminetetraacetic acid disodium salt (EDTA-2Na) on Enterococcus faecalis (E. faecalis) and Porphyromonas endodontalis (P. endodontalis). METHODS: E. faecalis and P. endodontalis were cultured and adjusted to 10(8) CFU/mL. Then 0.3, 0.5, 1, 2, 5, 10, 50, 100, 150 and 300 g/L of lysozyme were prepared with deionized water; and the lysozyme solutions were mixed with 0.5, 1.0, 2.0 g/L of EDTA-2Na, respectively. The bacteria and lysosome with/without EDTA-2Na interacted for 15 min, then water-soluble tetrazolium (WST) working solution was added and the activity of the bacteria was calculated by measuring optical densities at 450 nm and 630 nm with microplate spectrophotometer. RESULTS: Regarding the pure lysozyme from 0.5 g/L to 150 g/L, more E. faecalis and P. endodontalis were inhibited when the concentration of lysozyme was higher, especially for E. faecalis. There was synergistic effect of lysozyme with EDTA-2Na on antibacterial activity, which was related to the concentration of lysozyme. On E. faecalis, the antibacterial activity of lysozyme with EDTA-2Na was 1.2-3.7 folds than the pure lysozyme when the concentration of lysozyme was 0.5-50 g/L (P<0.05), and on P. endodontalis, the antibacterial activity of lysozyme with EDTA-2Na was 1.3-3.5 folds than the pure lysozyme when the concentration of lysozyme was 0.5-10 g/L (P<0.05). When the concentration of lysozyme was higher than 100 g/L, EDTA-2Na did not show synergistic effect on the antibacterial activity (P>0.05). CONCLUSION: For E. faecalis and P. endodontalis, a low concentration of lysozyme with EDTA-2Na showed significant synergistic antibacterial activity, while a high concentration of lysozyme with EDTA-2Na did not.

[Comparison of the incidences of apical root cracks after canal preparation with two nickel-titanium rotary systems: an in vitro study].

OBJECTIVE: To compare the incidences of root cracks after canal instrumentation with HyFlex CM system and the ProTaper Universal system. METHODS: Sixty mandibular incisors were mounted in resin blocks with simulated periodontal ligaments, and the apex was exposed. The control group of 20 teeth was not prepared, and the other 40 teeth were divided into 2 experimental groups (n=20). The 40 root canals of the experimental groups were instrumented using HyFlex CM and ProTaper Universal to the major apical foramen (AF). The horizontal sections 1 mm, 2 mm, and 3 mm from the apex were observed under an optical stereomicroscope at 25×magnification. The presence of cracks was noted. RESULTS: No cracks were found in the control teeth. Cracks were found in 1 of 20 (5%) teeth in HyFlex CM group, and 17 of 20 (85%) teeth in ProTaper Universal group. The difference between the two experimental groups was statistically significant (P<0.01). CONCLUSION: The HyFlex CM files caused fewer root cracks than the ProTaper Universal files during the root canal instrumentation.

Function of SREBP1 in the milk fat synthesis of dairy cow mammary epithelial cells.

Sterol regulatory element-binding proteins (SREBPs) belong to a family of nuclear transcription factors. The question of which is the most important positive regulator in milk fat synthesis in dairy cow mammary epithelial cells (DCMECs) between SREBPs or other nuclear transcription factors, such as peroxisome proliferator-activated receptor γ (PPARγ), remains a controversial one. Recent studies have found that mTORC1 (the mammalian target of rapamycin C1) regulates SREBP1 to promote fat synthesis. Thus far, however, the interaction between the SREBP1 and mTOR (the mammalian target of rapamycin) pathways in the regulation of milk fat synthesis remains poorly understood. This study aimed to identify the function of SREBP1 in milk fat synthesis and to characterize the relationship between SREBP1 and mTOR in DCMECs. The effects of SREBP1 overexpression and gene silencing on milk fat synthesis and the effects of stearic acid and serum on SREBP1 expression in the upregulation of milk fat synthesis were investigated in DCMECs using immunostaining, Western blotting, real-time quantitative PCR, lipid droplet staining, and detection kits for triglyceride content. SREBP1 was found to be a positive regulator of milk fat synthesis and was shown to be regulated by stearic acid and serum. These findings indicate that SREBP1 is the key positive regulator in milk fat synthesis.

[Surface roughness and staining stability of infiltrant resin for enamel white spot lesion].

OBJECTIVE: To investigate the influence factors on the surface roughness and staining susceptibility of infiltrant resin. METHODS: In the study, 30 human third molars were used, and each sample had three open enamel windows. The samples were randomly divided into three groups according to their different demineralized time. Each sample had at least one intact spot (A), one infiltrant resin spot (B) and one artificial white spot lesion (C). The surface roughness was tested before color staining for all the three spots of each specimen. The specimens were stored in a staining solution (coffee) for a period of 21 days, before and after which the color Commission Internationaled' Eclairage (CIE)L*a*b* was recorded for A, B and C spots. RESULTS: The B spot's surface roughness of each group was(0.15 ± 0.02)µm,(0.31 ± 0.03)µm and(0.40 ± 0.02)µm, respectively. And the C spot's surface roughness each was (1.08 ± 0.10)µm,(2.89 ± 0.13)µm and(3.41 ± 0.14)µm. The surface roughness of B and C of the three groups increased with demineralization time longer, and had significant difference for both B and C (P < 0.01). The ΔE of the three groups between A and B before staining had significant difference (P < 0.01). And the ΔE of group1 was less than 3.7, but the other two groups' more than 3.7. After staining, the ΔE of groups 1 and 2 was less than 3.7 but that of group 3 was more than 3.7. There were significant differences between groups 1 and 3, and also between groups 2 and 3(P < 0.01). CONCLUSION: The degree of the lesion's demineralization has effect on the surface roughness and color susceptibility of infiltrant resin. The increased surface roughness of infiltrant resin has positive effect on masking enamel white spots.

[Clinical evaluation of a two-step etch-and-rinse adhesive and a one-step self-etch adhesive in non-carious cervical lesion].

OBJECTIVE: To compare the clinical effectiveness of the two-step etch-and-rinse with the one-step self-etch adhesive in non-carious cervical lesions. METHODS: Fifty patients were selected, each with at least two wedge-shaped defects in the mouth. The paired defects were randomly bonded either with the two-step etch-and-rinse adhesive α or the one-step self-etch adhesive ß and then restored with resin composite. The treatment was carried out by one practitioner according to standard procedures. The evaluation was performed by another practitioner according to modified United States Public Health Service (USPHS) criteria at one week, six months and eighteen months after treatment. Chi-square test was used for statistical analysis. RESULTS: Fifty restorations were placed for each group. Forty-eight patients attended the six months recall, with two restorations loss for each group. Forty-four patients attended the eighteen months recall, with accumulative four restorations loss for adhesive α and six restorations loss for adhesive ß. The retention rate was 90.0% for group α and 86.4% for group ß. Marginal adaptation of three restorations in group α and five restorations in group ß were scored Bravo; while for marginal discoloration, two restorations in group α and three restorations in group ß were scored Bravo respectively. No secondary caries and post-operative sensitivity occurred for any of the restorations after eighteen months. No significant difference was detected between the groups for any evaluation criteria (P > 0.05). CONCLUSION: Within the observation period of this study, the two-step etch-and-rinse adhesive and the one-step self-etch adhesive showed similar clinical performance. The long term follow-up is still warranted.

[Influence of thermalcycling on bonding durability of self-etch adhesives with dentin].

OBJECTIVE: To investigate influence of thermalcycling on the bonding durability of two one-step products [Adper Prompt (AP) and G-bond (GB)] and one two-step self-etching adhesive [Clearfil SE bond (SE)] with dentin in vitro. METHODS: Forty-two extracted human molars were selected. The superficial dentin was exposed by grinding off the enamel. The teeth were randomly distributed into six groups with varied bonding protocols. The adhesives were applied to the dentin surface. Composite crowns were built up, then the samples were cut longitudinally into sticks with 1.0 mm×1.0 mm bonding area [for microtensile bond strength (MTBS) testing] or 1.0 mm thick slabs (for nanoleakage observation). Bonding performance was evaluated with or without thermalcyling. For the MTBS testing, the strength values were statistically analysed using One-Way ANOVA. Four slabs in each group were observed for nanoleakage by SEM with a backscattered electron detector. RESULTS: Thermalcycling procedures affected MTBS. In the two one-step groups, the MTBS decreased significantly (P<0.05) after thermalcycling [AP group from (19.06±1.50) MPa to (12.62±2.10) MPa; GB group from (17.75±1.10) MPa to (6.24±0.42)MPa]. But in SE groups, MTBS did not significantly affect [(45.80±2.97) MPa compared with(40.60±5.76) MPa]. As a whole, one-step self-etching adhesives showed lower MTBS than two-step bonding system after aging.For AP and GB, continuous nanoleakage appearance was notable and more obvious than for SE. CONCLUSION: Thermalcycling can affect the bonding performance of self-etch adhesives including decrease of bond strength and nanoleakage pattern. one-step self-etch adhesives showed more obvious change compared with their two-step counterparts.

[Notch activation delayed ageing of human dental pulp cells].

OBJECTIVE: To establish model of dental pulp cells with activated Notch signaling pathway, and investigate the effect of activating Notch signaling pathway on senescence of human dental pulp cells in vitro. METHODS: Human dental pulp cells were isolated, cultured as usual, and used from the 4(th) passage. The cells were divided into the activated group and the negative control group. In the activated group, the way of coating dishes with Jagged1 protein (10 mg/L) was used to activate Notch signaling pathway. The negative control group cells received no treatment. In the 4(th), 8(th), and 10(th) passages, the expression levels of the Notch signaling pathway downstream gene Hes1 were verified by real-time quantitative PCR (RT-qPCR). The cell changes after activating Notch signaling pathway were observed at three levels: (1) The cell morphology changes were observed through invert phase contrast microscope. The cell activity was detected with MTT assay. (2) The alkaline phosphatase (ALP) expression and its activity, and senescence-associated &bgr;-galatosidase (SA-ß-Gal) expression were observed with the kit. (3) The expression changes of senescence related genes were verified using RT-qPCR. The difference between the negative control group and the activated group was analyzed using student's t test. RESULTS: The expression level of the downstream gene Hes1 of Notch signaling pathway increased after coating the dishes with Jagged1 protein, indicating the establishment of the model of dental pulp cells with activated Notch signaling pathway. Compared with the negative control group, the aging cells of the activated group appeared relatively late. In the 8(th) and 10(th) passage, the cell activity increased. In the 10th passage, ALP activity increased, but SA-ß-Gal expression decreased. p16 gene expression decreased in each passage, and p53 gene expression decreased in the 8(th) and 10(th) passages. CONCLUSION: Jagged1 could activate Notch signaling pathway effectively. Through activating Notch signaling pathway, the dental pulp cells showed a trend of senescence delay at different levels, such as cell morphology, metabolic enzyme expressions and related gene expressions.

Goat mammary gland expression of Cecropin B to inhibit bacterial pathogens causing mastitis.

The antibacterial peptide Cecropin B (CB), isolated from the giant silk moth, has been shown to effectively eliminate bacteria. In this study, the effects of transgenic CB on dairy goat mammary epithelial cells (DGMECs) and dairy goat mammary gland were investigated. The DNA of CB from silkworm was amplified by reverse transcription PCR (RT-PCR) and then fused to the eukaryotic expression vector pECFP-C1. The recombinant plasmid pECFP-Cecropin B (pECFP-CB) was used for the transfection of DGMECs, and the expression of transgenic CB and the antibacterial activity of it were confirmed by western blot and agar diffusion reaction respectively. The stable DGMEC line transfected by pECFP-CB was obtained by screening with G418. In vivo experiment, pECFP-CB was injected into dairy goat mammary gland, and also the expression and antibacterial activity of transgenic CB were confirmed. Results of this study: transgenic CB can be expressed in DGMECs and dairy goat mammary gland, and inhibit the mastitis caused by Staphylococcus aureus.

SOCS3-mediated blockade reveals major contribution of JAK2/STAT5 signaling pathway to lactation and proliferation of dairy cow mammary epithelial cells in vitro.

Suppressor of cytokine signaling 3 (SOCS3) is a cytokine-induced negative feedback-loop regulator of cytokine signaling. More and more evidence has proved it to be an inhibitor of signal transducers and activators of transcription 5 (STAT5). Here, we used dairy cow mammary epithelial cells (DCMECs) to analyze the function of SOCS3 and the interaction between SOCS3 and STAT5a. The expression of SOCS3 was found in cytoplasm and nucleus of DCMECs by fluorescent immunostaining. Overexpression and inhibition of SOCS3 brought a remarkable milk protein synthesis change through the regulation of JAK2/STAT5a pathway activity, and SOCS3 expression also decreased SREBP-1c expression and fatty acid synthesis. Inhibited STAT5a activation correlated with reduced SOCS3 expression, which indicated that SOCS3 gene might be one of the targets of STAT5a activation, DCMECs treated with L-methionine (Met) resulted in a decrease of SOCS3 expression. SOCS3 could also decrease cell proliferation and viability by CASY-TT detection. Together, our findings indicate that SOCS3 acts as an inhibitor of JAK2/STAT5a pathway and disturbs fatty acid synthesis by decreasing SREBP-1c expression, which validates its involvement in both milk protein synthesis and fat synthesis. In aggregate, these results reveal that low SOCS3 expression is required for milk synthesis and proliferation of DCMECs in vitro.

[Effects of DAPT on proliferation of human dental pulp cells and Notch signaling pathway].

OBJECTIVE: To explore whether the γ-secretase inhibitor N-[N-(3,5-difluorophenacetyl-L-alanyl)]-S-phenylglycine t-butyl ester (DAPT) could inhibit Notch signaling pathway in human dental pulp cells, and its effects on the proliferation ability of the cells. METHODS: Human dental pulp cells were primarily cultured from healthy premolars or wisdom teeth extracted intactly. The γ-secretase inhibitor DAPT (5 µmol/L) was added to the culture medium from passage 4 to the end. The cells of passages 4, 8 and 10 were used as check points in this study. The Real-time RT-PCR and RT2 Profiler PCR Array were applied to analyze the expression changes of Notch signaling pathway downstream genes. And the methyl thiazolyl tetrazolium (MTT) method was used to test the proliferation ability of the cells. RESULTS: After DAPT was added, Hes1 gene expression level decreased significantly in the cells of passages 4, 8 and 10 as compared with that of the same passage cells in the control group. The relative gene expression ratio (experimental/control) decreased to 0.20 in the cells of passage 10, and the difference was significant (t=33.143,P=0.001). The PCR Array results of passage 10 dental pulp cells also showed a decline of Notch signaling pathway downstream genes Hey1 and NR4A2 in the experimental group as compared with the control group. The proliferation of passages 8, and 10 experimental cells were slowed down, and the difference was significant. CONCLUSION: Notch signaling pathway of human dental pulp cells could be inhibited by DAPT effectively. The proliferation of the cells was slowed down by the effect of DAPT, and the normal life cycle of the cells was affected.

[Relationship between volatile fatty acids and Porphyromonas gingivalis and Treponema denticola in gingival crevicular fluids of patients with aggressive periodontitis].

OBJECTIVE: Short chain fatty acids (SCFAs), such as succinic acid, acetic acid, propionic acid, butyric acid, etc. are metabolic product of putative periodontal pathogens, which play significant roles in periodontitis. The aim of this study was to analyze the relationship between Porphyromonas gingivalis (P. gingivalis), Treponema denticola (T. denticola), and the concentration of SCFAs in gingival crevicular fluid (GCF) of patients with aggressive periodontitis (AgP). METHODS: GCF was sampled from 4 sites per individual in 20 patients with AgP and 14 healthy controls. Concentrations of SCFAs, including succinic acid, acetic acid, propionic acid, butyric acid, and isovaleric acid in the supernant of GCF were analyzed by high performance capillary electrophoresis (HPCE), P. gingivalis and T. denticola in the deposit of the same GCF were detected by PCR with their electrophoretic band quantified. RESULTS: The concentrations of succinic acid, acetic acid, propionic acid, butyric acid, and isovaleric acid, the prevalence and PCR band quantity of P. gingivalis and T. denticola in GCF were all significantly higher in patients with AgP than that of healthy controls. In patients with AgP, butyric acid concentration was significantly higher in P. gingivalis positive sites than negative sites [2.87 (0.99, 4.36) mmol/L vs. 0.33 (0.00, 1.44) mmol/L, P<0.05], the concentrations of succinic acid, acetic acid, propionic acid, butyric acid, and isovaleric acid were positively correlated with PCR band quantity of P. gingivalis (r value was 0.334, 0.548, 0.411, 0.493, 0.273, respectively, P<0.05); the concentrations of SCFAs were significantly higher in T. denticola positive sites than negative sites: succinic acid, 1.67 (1.15, 2.11) mmol/L vs. 0.80 (0.48, 1.06) mmol/L; acetic acid, 31.95 (23.77, 43.13) mmol/L vs.12.51 (7.57, 15.69) mmol/L; propionic acid, 11.86 (6.55, 14.98) mmol/L vs. 2.82 (1.71, 7.03) mmol/L; butyric acid, 3.45 (2.41, 4.78) mmol/L vs. 0.54 (0.00, 1.56) mmol/L; isovaleric acid, 2.23 (1.05, 3.85) mmol/L vs. 0.62 (0.00, 2.33) mmol/L. The concentrations of succinic acid, acetic acid, propionic acid, butyric acid were positively correlated with PCR band quantity of T. denticola (r value was 0.443, 0.702, 0.625, 0.557, respectively, P<0.05). CONCLUSION: SCFAs concentrations reflect the quantity of P. gingivalis and T. denticola in patients with AgP, and may be an indicator to the disease progression in patients with AgP.

Safety and immunogenicity of a mosaic vaccine booster against Omicron and other SARS-CoV-2 variants: a randomized phase 2 trial.

An ongoing randomized, double-blind, controlled phase 2 trial was conducted to evaluate the safety and immunogenicity of a mosaic-type recombinant vaccine candidate, named NVSI-06-09, as a booster dose in subjects aged 18 years and older from the United Arab Emirates (UAE), who had administered two or three doses of inactivated vaccine BBIBP-CorV at least 6 months prior to enrollment. The participants were randomly assigned with 1:1 to receive a booster dose of NVSI-06-09 or BBIBP-CorV. The primary outcomes were immunogenicity and safety against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant, and the exploratory outcome was cross-immunogenicity against other circulating strains. Between May 25 and 30, 2022, 516 adults received booster vaccination with 260 in NVSI-06-09 group and 256 in BBIBP-CorV group. Interim results showed a similar safety profile between two booster groups, with low incidence of adverse reactions of grade 1 or 2. For immunogenicity, by day 14 post-booster, the fold rises in neutralizing antibody geometric mean titers (GMTs) from baseline elicited by NVSI-06-09 were remarkably higher than those by BBIBP-CorV against the prototype strain (19.67 vs 4.47-fold), Omicron BA.1.1 (42.35 vs 3.78-fold), BA.2 (25.09 vs 2.91-fold), BA.4 (22.42 vs 2.69-fold), and BA.5 variants (27.06 vs 4.73-fold). Similarly, the neutralizing GMTs boosted by NVSI-06-09 against Beta and Delta variants were also 6.60-fold and 7.17-fold higher than those by BBIBP-CorV. Our findings indicated that a booster dose of NVSI-06-09 was well-tolerated and elicited broad-spectrum neutralizing responses against divergent SARS-CoV-2 variants, including Omicron and its sub-lineages.

Proteomic and functional analyses reveal MAPK1 regulates milk protein synthesis.

L-Lysine (L-Lys) is an essential amino acid that plays fundamental roles in protein synthesis. Many nuclear phosphorylated proteins such as Stat5 and mTOR regulate milk protein synthesis. However, the details of milk protein synthesis control at the transcript and translational levels are not well known. In this current study, a two-dimensional gel electrophoresis (2-DE)/MS-based proteomic technology was used to identify phosphoproteins responsible for milk protein synthesis in dairy cow mammary epithelial cells (DCMECs). The effect of L-Lys on DCMECs was analyzed by CASY technology and reversed phase high performance liquid chromatography (RP-HPLC). The results showed that cell proliferation ability and ß-casein expression were enhanced in DCMECs treated with L-Lys. By phosphoproteomics analysis, six proteins, including MAPK1, were identified up-expressed in DCMECs treated with 1.2 mM L-Lys for 24 h, and were verified by quantitative real-time PCR (qRT-PCR) and western blot. Overexpression and siRNA inhibition of MAPK1 experiments showed that MAPK1 upregulated milk protein synthesis through Stat5 and mTOR pathway. These findings that MAPK1 involves in regulation of milk synthesis shed new insights for understanding the mechanisms of milk protein synthesis.

MiR-15a decreases bovine mammary epithelial cell viability and lactation and regulates growth hormone receptor expression.

MicroRNAs (miRNAs) are a class of small non-coding RNAs that regulate the expression of target genes at the post-transcriptional level by transcript degradation or translational inhibition. The role of bta-miR-15a in bovine mammary gland hasn't been reported. Using miRNAs prediction software, GHR gene was predicted to be a potential target of bta-miR-15a. In this study, bovine mammary epithelial cell line was used as an in vitro cell model to address the function of bta-miR-15a on bovine mammary epithelial cells. The expression changes of bta-miR-15a and Ghr after bta-miR-15a transfection were detected by qRT-PCR; the expression of GHR protein and casein was detected by western blotting. To determine whether bta-miR-15a can affect cell viability, cells were examined using an electronic Coulter counter (CASY-TT). In conclusion, bta-miR-15a inhibited the expression of casein of bovine mammary epithelial cells, and cell number and viability were reduced by bta-miR-15a expression. Bta-miR-15a inhibited the viability of mammary epithelial cells as well as the expression of GHR mRNA and protein level, therefore suggesting that bta-miR-15a may play an important role in mammary gland physiology.