MicroRNA-21 promotes wound healing via the Smad7-Smad2/3-Elastin pathway.

Wound healing is regulated by a complex network of cells, molecules, and cytokines, as well as microRNAs (miRNAs). miRNAs were confirmed to influence the wound healing process, and miR-21, an important member of the miRNA family, was also shown to regulate wound healing. The aim of the present study was to investigate the role of miR-21 in the wound healing process and the possible underlying cell signaling pathways. We isolated GMSCs from WT and miR-21-KO mouse gingiva. Flow cytometric analysis and immunocytofluorescense staining were used to identify the GMSCs acquired from WT and miR-21-KO mice. RT-PCR, western blot analysis and immunohistofluorescence staining were performed to examine the expression of extracellular matrix components and key proteins of cell signaling pathways. TargetScan and pmiR-RB-REPORT vectors were used to verify that Smad7 was a direct target of miR-21. Compared to WT mice, miR-21-KO mice showed slower wound healing. RT-PCR and western blot analysis indicated that Elastin expression was downregulated in miR-21-deficient samples. We confirmed that Smad7 was a direct target of miR-21. miR-21 knockout resulted in increased expression of Smad7 and impaired phosphorylation of the Smad2/3 complex. The expression of the Smad7-Smad2/3-Elastin axis in palate tissues sections acquired from WT and miR-21-KO mice showed the same trend. Based on all these results, we demonstrated that miR-21 promoted the wound healing process via the Smad7-Smad2/3-Elastin pathway.

SMAD4-mediated WNT signaling controls the fate of cranial neural crest cells during tooth morphogenesis.

TGFß/BMP signaling regulates the fate of multipotential cranial neural crest (CNC) cells during tooth and jawbone formation as these cells differentiate into odontoblasts and osteoblasts, respectively. The functional significance of SMAD4, the common mediator of TGFß/BMP signaling, in regulating the fate of CNC cells remains unclear. In this study, we investigated the mechanism of SMAD4 in regulating the fate of CNC-derived dental mesenchymal cells through tissue-specific inactivation of Smad4. Ablation of Smad4 results in defects in odontoblast differentiation and dentin formation. Moreover, ectopic bone-like structures replaced normal dentin in the teeth of Osr2-IresCre;Smad4(fl/fl) mice. Despite the lack of dentin, enamel formation appeared unaffected in Osr2-IresCre;Smad4(fl/fl) mice, challenging the paradigm that the initiation of enamel development depends on normal dentin formation. At the molecular level, loss of Smad4 results in downregulation of the WNT pathway inhibitors Dkk1 and Sfrp1 and in the upregulation of canonical WNT signaling, including increased ß-catenin activity. More importantly, inhibition of the upregulated canonical WNT pathway in Osr2-IresCre;Smad4(fl/fl) dental mesenchyme in vitro partially rescued the CNC cell fate change. Taken together, our study demonstrates that SMAD4 plays a crucial role in regulating the interplay between TGFß/BMP and WNT signaling to ensure the proper CNC cell fate decision during organogenesis.

Literature review and prospect on oral cognition and disease diagnosis and treatment between Han and Tang dynasties.

Chinese medicine entered a significant period from foundation to maturity between Han and Tang dynasties when the Chinese traditional stomatology was a key stage. Sorting and analysis of existing literature and research outcomes have showed that current research on stomatology between Han and Tang dynasties focuses on oral physiology, pathology, diagnosis and treatment, and health care. It also involves stomatology history and explanation of termino-logies related to mouth and teeth recorded in medical books, use of simple methods, and thinking with citation and analysis of literature simply listed and reasoning preliminarily deducted. From the macro perspective, current research has not unveiled the whole picture of stomatology between the two dynasties and left a series of key issues unresolved. Thus, new methods should be developed and employed to carry out medical research on stomatology between Han and Tang dynasties given that is has a prosperous future.

Progress and perspectives on cell-free regenerative endodontic therapy.

Although teeth with pulpitis/apical periodontitis are saved after successful endodontic therapy, they are devitalized and susceptible to reinfections and fractures. The development of biology-based approaches for dental pulp regeneration or repair is possible today because of recent advances in tissue engineering and biomaterials. Cell-free regenerative endodontic therapy offers a promising strategy for the treatment of necrotic immature permanent teeth in children and adolescents. However, studies are underway to determine whether this procedure can be applied to mature teeth.

Safety and efficacy assessment of allogeneic human dental pulp stem cells to treat patients with severe COVID-19: structured summary of a study protocol for a randomized controlled trial (Phase I / II).

OBJECTIVES: To assess the safety and therapeutic effects of allogeneic human dental pulp stem cells (DPSCs) in treating severe pneumonia caused by COVID-19. TRIAL DESIGN: This is a single centre, two arm ratio 1:1, triple blinded, randomized, placebo-controlled, parallel group, clinical trial. PARTICIPANTS: Twenty serious COVID-19 cases will be enrolled in the trial from April 6th to December 31st 2020. INCLUSION CRITERIA: hospitalised patients at Renmin Hospital of Wuhan University satisfy all criteria as below: 1)Adults aged 18-65 years;2)Voluntarily participate in this clinical trial and sign the "informed consent form" or have consent from a legal representative.3)Diagnosed with severe pneumonia of COVID-19: nucleic acid test SARS-CoV-2 positive; respiratory distress (respiratory rate > 30 times / min); hypoxia (resting oxygen saturation < 93% or arterial partial pressure of oxygen / oxygen concentration < 300 mmHg).4)COVID-19 featured lung lesions in chest X-ray image. EXCLUSION CRITERIA: Patients will be excluded from the study if they meet any of the following criteria. 1.Patients have received other experimental treatment for COVID-19 within the last 30 days;2.Patients have severe liver condition (e.g., Child Pugh score >=C or AST> 5 times of the upper limit);3.Patients with severe renal insufficiency (estimated glomerular filtration rate <=30mL / min/1.73 m2) or patients receiving continuous renal replacement therapy, hemodialysis, peritoneal dialysis;4.Patients who are co-infected with HIV, hepatitis B, tuberculosis, influenza virus, adenovirus or other respiratory infection viruses;5.Female patients who have no sexual protection in the last 30 days prior to the screening assessment;6.Pregnant or lactating women or women using estrogen contraception;7.Patients who are planning to become pregnant during the study period or within 6 months after the end of the study period;8.Other conditions that the researchers consider not suitable for participating in this clinical trial. INTERVENTION AND COMPARATOR: There will be two study groups: experimental and control. Both will receive all necessary routine treatment for COVID-19. The experimental group will receive an intravenous injection of dental pulp stem cells suspension (3.0x107 human DPSCs in 30ml saline solution) on day 1, 4 and 7; The control group will receive an equal amount of saline (placebo) on the same days. Clinical and laboratory observations will be performed for analysis during a period of 28 days for each case since the commencement of the study. MAIN OUTCOMES: 1. Primary outcome The primary outcome is Time To Clinical Improvement (TTCI). By definition, TTCI is the time (days) it takes to downgrade two levels from the following six ordered grades [(grade 1) discharge to (grade 6) death] in the clinical state of admission to the start of study treatments (hDPSCs or placebo). Six grades of ordered variables: GradeDescriptionGrade 1:Discharged of patient;Grade 2:Hospitalized without oxygen supplement;Grade 3:Hospitalized, oxygen supplement is required, but NIV / HFNC is not required;Grade 4:Hospitalized in intensive care unit, and NIV / HFNC treatment is required;Grade 5:Hospitalized in intensive care unit, requiring ECMO and/or IMV;Grade 6:Death. ABBREVIATIONS: NIV, non-invasive mechanical ventilation; HFNC, high-flow nasal catheter; IMV, invasive mechanical ventilation. 2. Secondary outcomes 2.1 vital signs: heart rate, blood pressure (systolic blood pressure, diastolic blood pressure). During the screening period, hospitalization every day (additional time points of D1, D4, D7 30min before injection, 2h ± 30min, 24h ± 30min after the injection) and follow-up period D90 ± 3 days. 2.2 Laboratory examinations: during the screening period, 30 minutes before D1, D4, D7 infusion, 2h ± 30min, 24h ± 30min after the end of infusion, D10, D14, D28 during hospitalization or discharge day and follow-up period D90 ± 3 days. 2.3 Blood routine: white blood cells, neutrophils, lymphocytes, monocytes, eosinophils, basophils, neutrophils, lymphocytes, monocytes, eosinophils Acidic granulocyte count, basophil count, red blood cell, hemoglobin, hematocrit, average volume of red blood cells, average red blood cell Hb content, average red blood cell Hb concentration, RDW standard deviation, RDW coefficient of variation, platelet count, platelet specific platelet average Volume, platelet distribution width,% of large platelets; 2.4 Liver and kidney function tests: alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, γ-glutamyl transferase, prealbumin, total protein, albumin, globulin, white / globule ratio , Total bilirubin, direct bilirubin, cholinesterase, urea, creatinine, total carbon dioxide, uric acid glucose, potassium, sodium, chlorine, calcium, corrected calcium, magnesium, phosphorus, calcium and phosphorus product, anion gap, penetration Pressure, total cholesterol, triacylglycerol, high density lipoprotein cholesterol, Low density lipoprotein cholesterol, lipoprotein a, creatine kinase, lactate dehydrogenase, estimated glomerular filtration rate. 2.5 Inflammation indicators: hypersensitive C-reactive protein, serum amyloid (SAA); 2.6 Infectious disease testing: Hepatitis B (HBsAg, HBsAb, HBeAg, HBeAb, HBcAb), Hepatitis C (Anti-HCV), AIDS (HIVcombin), syphilis (Anti-TP), cytomegalovirus CMV-IgM, cytomegalovirus CMV-IgG; only during the screening period and follow-up period D90 ± 3. 2.7 Immunological testing: Collect peripheral blood to detect the phenotype of T lymphocyte, B lymphocyte, natural killer cell, Macrophage and neutrophil by using flow cytometry. Collect peripheral blood to detect the gene profile of mononuclear cells by using single-cell analyses. Collect peripheral blood serum to detect various immunoglobulin changes: IgA, IgG, IgM, total IgE; Collect peripheral blood serum to explore the changes of cytokines, Th1 cytokines (IL-1 ß, IL-2, TNF-a, ITN-γ), Th2 cytokines (IL-4, IL-6, IL -10). 2.8 Pregnancy test: blood ß-HCG, female subjects before menopause are examined during the screening period and follow-up period D90 ± 3. 2.9 Urine routine: color, clarity, urine sugar, bilirubin, ketone bodies, specific gravity, pH, urobilinogen, nitrite, protein, occult blood, leukocyte enzymes, red blood cells, white blood cells, epithelial cells, non-squamous epithelial cells , Transparent cast, pathological cast, crystal, fungus; 2.10 Stool Routine: color, traits, white blood cells, red blood cells, fat globules, eggs of parasites, fungi, occult blood (chemical method), occult blood (immune method), transferrin (2h ± 30min after the injection and not detected after discharge). RANDOMIZATION: Block randomization method will be applied by computer to allocate the participants into experimental and control groups. The random ratio is 1:1. BLINDING (MASKING): Participants, outcomes assessors and investigators (including personnel in laboratory and imaging department who issue the sample report or image observations) will be blinded. Injections of cell suspension and saline will be coded in accordance with the patient's randomisation group. The blind strategy is kept by an investigator who does not deliver the medical care or assess primary outcome results. NUMBERS TO BE RANDOMIZED (SAMPLE SIZE): Twenty participants will be randomized to the experimental and control groups (10 per group). TRIAL STATUS: Protocol version number, hDPSC-CoVID-2019-02-2020 Version 2.0, March 13, 2020. Patients screening commenced on 16th April and an estimated date of the recruitment of the final participants will be around end of July. . TRIAL REGISTRATION: Registration: World Health Organization Trial Registry: ChiCTR2000031319; March 27,2020. ClinicalTrials.gov Identifier: NCT04336254; April 7, 2020 Other Study ID Numbers: hDPSC-CoVID-2019-02-2020 FULL PROTOCOL: The full protocol is attached as an additional file, accessible from the Trials website (Additional file 1). In the interest in expediting dissemination of this material, the familiar formatting has been eliminated; this Letter serves as a summary of the key elements of the full protocol.

[Increase of saliva nitrate and nitrite level in patients with oral candidiasis].

OBJECTIVE: To observe the alterations of saliva nitrate and nitrite level in patients with oral candidiasis. METHODS: Parotid saliva and whole saliva were collected from 33 patients and 34 healthy volunteers. Concentrations of nitrate and nitrite in saliva were determined by high-performance liquid chromatography. Follow-up observation was performed on 10 patients after treatment. The data were statistically analyzed with independent-samples t test or paired-samples t test at alpha = 0.05. RESULTS: There was significant increase of the concentrations and secretion rate of parotid saliva nitrate in patient group as compared with controls: (49.70 +/- 0.50) vs (21.51 +/- 0.60) mg/L (t = 2.692, P = 0.009) and (27.71 +/- 0.50) vs (12.55 +/- 0.60) microg/min (t = 2.554, P = 0.013), respectively. Significantly increased concentrations and secretion rate of nitrate and nitrite [nitrate: (6.46 +/- 0.94) vs (1.11 +/- 0.70) mg/L (t = 3.792, P = 0.000); nitrite: (8.48 +/- 0.58) vs (3.39 +/- 0.53) mg/L (t = 2.888, P = 0.005); nitrate secretion rate: (10.57 +/- 0.91) vs (2.10 +/- 0.74) microg/min (t = 3.464, P= 0.001); nitrite secretion rate: (13.91 +/- 0.55) vs (6.42 +/- 0.58) microg/min (t = 2.397, P = 0.020)] were revealed in whole saliva of patients group. Significantly decreased nitrate and nitrite levels were also observed in patients after treatment, especially the changes of parotid saliva nitrate secretion rate [(37.50 +/- 0.50) vs (14.34 +/- 0.64) microg/min (t = 3.142, P = 0.012)], whole saliva nitrate [(14.29 +/- 1.01) vs (2.59 +/- 1.03) mg/L (t = 3.475, P = 0.007)] and whole saliva nitrate secretion rate [(25.97 +/- 0.93) vs (4.12 +/- 1.00) microg/min (t = 3.922, P = 0.003)]. CONCLUSION: The present study revealed the significant increase of salivary nitrate and nitrite level in patients with oral candidiasis is considered to be associated with the host defense reaction.

Aspirin inhibits LPS-induced macrophage activation via the NF-κB pathway.

Aspirin (acetylsalicylic acid, ASA) has been shown to improve bone marrow mesenchymal stem cell-based calvarial bone regeneration by promoting osteogenesis and inhibiting osteoclastogenesis. However, it remains unknown whether aspirin influences other immune cells during bone formation. In the present study, we investigated whether ASA treatment influenced macrophage activation during the LPS inducement. We found that ASA could downregulate the expressions of iNOS and TNF-α both in mouse peritoneum macrophages and RAW264.7 cells induced by LPS via the IκK/IκB/NF-κB pathway and a COX2/PGE2/EP2/NF-κB feedback loop, without affecting the expressions of FIZZ/YM-1/ARG1 induced by IL-4. Furthermore, we created a rat mandibular bone defect model and showed that ASA treatment improved bone regeneration by inhibiting LPS-induced macrophage activation in the early stages of inflammation. Taken together, our results indicated that ASA treatment was a feasible strategy for improving bone regeneration, particularly in inflammatory conditions.

Toll-like receptor 4 polymorphism is associated with increased susceptibility to chronic obstructive pulmonary disease in Han Chinese patients with chronic periodontitis.

The aim of the present study was to evaluate the association of single nucleotide polymorphisms (SNPs) in toll-like receptor 4 (TLR4) with chronic obstructive pulmonary disease (COPD) in Han Chinese patients with chronic periodontitis (CP). Six candidate SNPs of TLR4-rs10759930, rs10983755, rs11536879, rs1927907, rs11536889 and rs7873784-and 18 haplotype-tagging SNPs (tagSNPs) were genotyped in 339 patients with chronic periodontitis only (CP group), and 373 CP patients with COPD (CP with COPD group). The genotype distribution and allele frequencies of TLR4 rs1927907 among the CP (AA: 26, 8.5%, AG: 109, 35.5%, GG: 172, 56.0%) and CP with COPD (AA: 41, 12.0%, AG: 143, 41.7%, GG: 159, 46.4%) groups were significantly different (P = 0.039). After adjusting for age, sex, smoking status, and oral hygiene habits, CP patients carrying the AG polymorphism in TLR4 rs1927907 were found to be more susceptible to concomitant COPD than those carrying the GG genotype (P = 0.005, OR = 1.94, 95% CI for OR: 1.22-3.03). In conclusion, TLR4 gene polymorphism plays a role in the common pathophysiology of CP and COPD, indicating that CP patients with TLR4 gene rs1927907 polymorphism may be more susceptible to COPD.(J Oral Sci 58, 555-560, 2016).

Morphological characteristics of submandibular glands of miniature pig.

BACKGROUND: Miniature pig (minipig) is increasingly used as a large animal model for a variety of biomedical studies. Little information is available in the literature on anatomy, histology and sialograghy of the submandibular gland of the minipig. The purpose of this study was to characterize the morphology of a miniature pig's (minipig) submandibular gland as a large animal model for further biomedical studies. METHODS: Five minipigs were subjected to sialographic, anatomic, histologic, histochemical and ultrastructural evaluations for submandibular glands. RESULTS: Sialograms showed a long, horizontal main excretory duct and a pear-shaped gland located inferoposterior to the angle of the mandible. The submandibular glands lied superficial to the suprahyoid, and infrahyoid muscle groups, and were covered by the inferior portion of the parotid gland. The submandibular glands were characterized by a mixed parenchyma of mucous and serous secretory acini. Alcian blue (AB) staining and periodic acid-Schiff (PAS) reactions demonstrated that minipig submandibular glands synthesized and secreted acid mucous substances by serous cells and polysaccharide, and neutral mucous substances, by mucous cells. CONCLUSION: The submandibular gland of the minipig is considered a useful large salivary gland animal model for biomedical studies.

[Tooth regeneration--dream to reality].

Tooth or dentition missing compromises human health physically and psychiatrically. Although several prosthesis methods are used to restore tooth loss, these restorations are still non-biological methods. It is a dream for human being to regenerate a real tooth for hundreds years. There are two ways to regenerate the tooth. One is application of conventional tissue engineering techniques including seed cells and scaffold. The other is regeneration tooth using dental epithelium and dental mesenchymal cells based on the knowledge of tooth initiation and development. Marked progress has been achieved in these two ways, while there is still a long way to go. Recently a new concept has been proposed for regeneration of a biological tooth root based on tooth-related stem cells and tissue engineering technique. A biological tooth root has been regenerated in swine. It may be a valuable method for restoration of tooth loss before successful whole tooth regeneration. A latest research showed that a subpopulation in bone marrow cells can give rise to ameloblast-like cells when mixed with embryonic epithelium and reassociation with integrated mesenchyme, which may provide a new seed cell source for tooth regeneration.

[Alterations of salivary flow rate and oral main pathogens in miniature pig with bilateral parotid gland atrophy].

OBJECTIVE: To observe the effect of bilateral parotid gland atrophy on the whole saliva flow rate and the growth of main oral pathogens in different sites of oral cavity. METHODS: Ten healthy miniature pigs were divided into two groups. The parotid glands of test group (n = 5) were bilaterally ablated by methyl violet. Another healthy five miniature pigs served as the control group. Whole saliva was collected and the whole saliva flow rate detected in both groups at 12 and 24 months respectively after parotid atrophy. The total numbers of oral main pathogens in the first molar, cuspid sub-gingival bacteria plaque and whole saliva were also detected. RESULTS: The whole saliva flow rate was significantly decreased at both 12 and 24 months respectively after atrophy of bilateral parotid gland in miniature pig. Pathogens including Streptococcus mutans, Porphyromonas gingivalis and Fusobacterium nucleatum in different sites oral cavity were increased after bilateral parotid gland atrophy. CONCLUSIONS: Bilateral ablation of the parotid glands led to a significant decrease of whole saliva flow rate. The total numbers of main oral pathogens were increased in different sites of oral cavity.

[Tooth tissue engineering: from cells to organ, an odyssey far from finished].

Tooth tissue engineering is an emerging biotechnique that will provide replacemental teeth for patients suffering from different diseases causing tooth loss. Although some attempts have been tried to generate whole tooth both in vivo and in vitro, the lack of the knowledge for tooth initiation and development, as well as for tooth shape controlling mechanisms greatly impede the progress of this technique. This article reviewed and discussed some recent findings in tooth tissue engineering related to the cell resource, the concept of reconstruction and regeneration, the application of artificial scaffolds, together with the methods of organ culture and implantation.

[Sialographic, sialoendoscopic and irrigation fluid study in chronic obstructive parotitis].

OBJECTIVE: To study the sialographic changes and to compare the changes with sialoendoscopic and irrigation fluid findings in chronic obstructive parotitis (COP). METHODS: This study involved 27 patients with a long history of parotid swelling. All patients were examined by X-ray, sialography, and were diagnosed as COP without sialolithiasis. Sialoendoscopy was used to observe the ductal system and irrigation treatment performed. The irrigated liquid was centrifuged and the deposits of fluid were stained and observed under microscopy. The sialographic changes were classified as previous studies and compared with sialoendoscopic and irrigation fluid findings. RESULTS: The sialographic changes of COP in 27 patients included 9 cases with type I, 5 cases with type II, 9 cases with type III and 3 cases with type IV changes, 1 case was normal. Marked obstructive factors such as stricture of ductal system were revealed in 21 cases on the sialogram. Sialoendoscopic examination showed that the ductal system was filled with fiber-like substances and hyperaemia of ductal wall in all cases. While few and thin fiber-like substances were found in the COP with sialographic type I and type II changes, many thick wadding or mass fiber-like substances were revealed in COP with sialographic type III and IV changes. Microstones were found in 2 COP with sialographic type III changes which were stained and identified by microscopy. Foreign body (drug bar) was found in one COP with sialographic type I changes with sialoendoscopy. Irrigation fluid examination showed fiber-like substance was composed of desquamative duct epithelial cells, neutrophil, lymphocytes, acidophile. Some epithelial cells were found in two microliths. CONCLUSIONS: The pathological basis of fiber-like substance on sialoendoscopy is desquamative duct epithelial cells. Fiber-like substance in the lumen of ductal system is considered as one of the obstructive factors in COP. Sialoendoscopic findings is related to sialographic changes.

[Effect of irradiation on tooth hard tissue and its resistance to acid].

OBJECTIVE: To study the effect of irradiation on the susceptibility of radiation caries. METHODS: The structures of 56 teeth enamel and dentin of 63 roots were observed using SEM and the collagen fibre and the resistance to the acid were also investigated after irradiation of 30 Gy, 50 Gy and 70 Gy. RESULTS: The enamel structure changes were found after irradiation with different doses. The significant difference was found in the enamel changes between high or middle dose group and low dose group or control. The dentin morphology changed, some collagen fibre vanished and resistance to acid was reduced after irradiation with 50 Gy and 70 Gy. CONCLUSIONS: The radiation reduced the resistance of teeth to the acid and increased the caries susceptibility.

[The study of the relationship of malodor and microbial composition of interdental and subgingival plaques in periodontitis patients].

OBJECTIVE: The study is to investigate the microbial composition of interdental and subgingival plaques of periodontitis patients with or without malodor, to explore the relationships between periodontitis and oral malodor. METHODS: 20 patients of periodontitis with malodor were chosen from 210 patients of periodontitis, and the clinical parameter of plaque index (PLI), gingival bleeding index (GBI) and probing depth (PD) were measured and compared with the control group which had periodontal disease without malodor. During the experiment, the interdental and subgingival microbial samples in both groups were collected and sent to anaerobic culture for 48 hrs, then the total CFU/ml of each sample were counted, and each type of bacteria was separated and identified. All of the data were analyzed by using the statistical software SPSS 10.0. RESULTS: (1) There were no satistical differences on PLI, GBI, PD between experimental group and control group. (2) The percents of leptospira in both interdental and subgingival plaques of test group were significantly higher than that of the control group (P < 0.01). (3) Either the interdental or in subgingival plaques, the count results of CFU/ml were similar in both groups (P > 0.05). (4) The proportions of malodor producing anaerobic bacteria in interdental gingival plaque, such as P. gingivalis and Veillonelia, were singnificantly different between test group and control group. CONCLUSION: The proportions of VSC's producing anaerobic bacteria in interdental gingival plaque may be play the significant roles in oral malodor. Further studies should be taken to elucidate the relationship between malodor and periodontitis.