Improving Blood Monocyte Energy Metabolism Enhances Its Ability to Phagocytose Amyloid-ß and Prevents Alzheimer's Disease-Type Pathology and Cognitive Deficits.

Deficiencies in the clearance of peripheral amyloid ß (Aß) play a crucial role in the progression of Alzheimer's disease (AD). Previous studies have shown that the ability of blood monocytes to phagocytose Aß is decreased in AD. However, the exact mechanism of Aß clearance dysfunction in AD monocytes remains unclear. In the present study, we found that blood monocytes in AD mice exhibited decreases in energy metabolism, which was accompanied by cellular senescence, a senescence-associated secretory phenotype, and dysfunctional phagocytosis of Aß. Improving energy metabolism rejuvenated monocytes and enhanced their ability to phagocytose Aß in vivo and in vitro. Moreover, enhancing blood monocyte Aß phagocytosis by improving energy metabolism alleviated brain Aß deposition and neuroinflammation and eventually improved cognitive function in AD mice. This study reveals a new mechanism of impaired Aß phagocytosis in monocytes and provides evidence that restoring their energy metabolism may be a novel therapeutic strategy for AD.

Associations of Blood and Cerebrospinal Fluid Aß and tau Levels with Renal Function.

Amyloid ß (Aß) and tau play pivotal roles in the pathogenesis of Alzheimer's disease (AD). Previous studies have shown that brain-derived Aß and tau can be cleared through transport into the periphery, and the kidneys may be vital organs involved in the clearance of Aß and tau. However, the effects of deficiency in the clearance of Aß and tau by the kidneys on brain AD-type pathologies in humans remain largely unknown. In this study, we first recruited 41 patients with chronic kidney disease (CKD) and 40 age- and sex-matched controls with normal renal function to analyze the associations of the estimated glomerular filtration rate (eGFR) with plasma Aß and tau levels. To analyze the associations of eGFR with cerebrospinal fluid (CSF) AD biomarkers, we recruited 42 cognitively normal CKD patients and 150 cognitively normal controls with CSF samples. Compared with controls with normal renal function, CKD patients had higher plasma levels of Aß40, Aß42 and total tau (T-tau), lower CSF levels of Aß40 and Aß42 and higher levels of CSF T-tau/Aß42 and phosphorylated tau (P-tau)/Aß42. Plasma Aß40, Aß42, and T-tau levels were negatively correlated with eGFR. In addition, eGFR was negatively correlated with CSF levels of T-tau, T-tau/Aß42, and P-tau/Aß42 but positively correlated with Mini-Mental State Examination (MMSE) scores. Thus, this study showed that the decline in renal function was correlated with abnormal AD biomarkers and cognitive decline, which provides human evidence that renal function may be involved in the pathogenesis of AD.

Enhanced conservation of vital pulp and apical tissues by the application of crown rotation surgery for inversely impacted central incisors: A follow-up analysis of two patients over 4 years.

AIM: To describe a novel surgical method (crown rotation surgery) to manage inversely impacted central incisors with immature roots. METHODOLOGY: Two young patients each presented with an inversely impacted maxillary central incisor. To protect the apical tissues, the two impacted incisors were rotated downwards to a relatively normal position without extraction from their bony sockets. RESULTS: After crown rotation surgery, spontaneous eruption, continuous root development, and periodontal healing of the rotated incisors were observed. The pulp retained vitality and blood flow was normal. Moreover, there were no obvious signs of pulp canal obliteration (PCO), as indicated by Cone Beam Computed Tomography (CBCT) imaging. CONCLUSIONS: By optimising protection of the vital pulp and apical tissues, crown rotation surgery represents a minimally invasive, conservative, and practical surgical technique for treating inversely impacted incisors with developing roots. In contrast to existing surgical methods, crown rotation surgery may avoid certain complications, including PCO and abnormal or arrested root development. KEY LEARNING POINTS: By optimizing protection of the vital pulp and apical tissues, crown rotation surgery represents a minimally invasive, conservative and practical surgical technique for treating inversely impacted incisors with developing roots. In contrast to existing surgical methods, crown rotation surgery may avoid certain complications, including PCO and abnormal or arrested root development.

Cucurbitacin B exhibits antitumor effects on CD133+ HepG2 liver cancer stem cells by inhibiting JAK2/STAT3 signaling pathway.

Cancer stem cells (CSCs), a crucial cancer cell subpopulation, possess stemness phenotypic characteristics. Cucurbitacin B (CuB), a tetracyclic triterpenoid isolated from Cucurbitaceae, exerts widely pharmacological activities in many diseases. The aim of this study was to enrich, identify liver CSCs and investigate antitumor effects of CuB as well as explore the underlying molecular mechanisms in these liver CSCs. HepG2 cell lines were used for the enrichment of liver CSCs by serum-free medium culture and magnetic-activated cell sorting. The CSC characteristics were analyzed by immunofluorescent staining, sphere-forming, western blot and xenograft tumorigenicity assay. CuB' antitumor effects and underlying molecular mechanism were measured by cell counting kit-8, colony formation, sphere-forming, cell cycle, xenograft and western blot assay. Our results showed that we could enrich 97.29% CD133+ HepG2 cells, which possessed CSC characteristics including re-renewal capacity, proliferative ability, sorafenib resistance, overexpressed stemness-related molecules and enhanced tumorigenic potential. Furthermore, we also found that CuB inhibited cell viability, sphere formation, colony formation and arrested cell cycle at G2/M phase as well as sensitized CD133+ HepG2 cells to sorafenib in vitro and in vivo. Western blot assay indicated that CuB inhibited expression levels of cyclin B1, CDK1, CD133, p-JAK2 and p-STAT3. In conclusion, our findings indicated that CuB could exhibit antitumor effects on CD133+ HepG2 CSCs by inhibiting the Janus kinase 2/signal transducers and activators of transcription-3 signaling pathway, expanding basic and preclinical investigations on liver CSCs.

In situ rotation surgery for correction of growing, inversely impacted maxillary central incisors.

Treatment of an impacted incisor with a dilacerated root is challenging for clinicians because of the position of the impacted incisor, the abnormality of the root, unfavorable prognosis, and, especially, the long treatment duration. We report on 2 young patients who had inversely impacted maxillary central incisors with developing labially dilacerated roots. Both patients were treated by a novel surgical approach, in situ rotation, by which the crowns of the inversely impacted incisors were carefully rotated to a relatively normal position, whereas the apical location remained relatively unchanged. About 2 weeks after surgery, spontaneous eruption of the treated incisors was observed. Three months later, the postoperative central incisors were further aligned into the maxillary arch with a fixed orthodontic appliance. Follow-up visits 2 or 3 years after surgery indicated that the positions of the dilacerated incisors maintained stability with good gingival esthetics, and the pulpal vitality was favorable. The roots grew further in a relatively normal direction of the incisor's longitudinal axis, which was different from the initial curvature angle. Moreover, with the in situ rotation surgery, treatment time was greatly reduced and resulted in a favorable prognosis compared with conventional treatment.

Sorafenib and CuB exert synergistic antitumor effects against hepatocellular carcinoma cells via inhibition of STAT3 phosphorylation.

Sorafenib, the first-line agent for treatment of advanced hepatocellular carcinoma (HCC), improves median overall survival by approximately 3 months. In the present study, we investigated whether sorafenib combined with cucurbitacin B (CuB), a natural tetracyclic triterpenoid isolated from Cucurbitaceae, exerts enhanced antitumor effects against HCC. Cell viability and colony formation ability were detected by cell-counting kit-8 and colony formation assays. Cell cycle and apoptosis were analyzed by flow cytometry. Protein expression was detected by western blotting. HepG2 xenografts in nude mice were used to evaluate in vivo antitumor effects. We report that sorafenib and CuB exhibited synergistic effects on cellular proliferation inhibition and cell apoptosis induction, but not on cell cycle arrest. Furthermore, combination treatment enhanced levels of cleaved caspase 3 and cleaved caspase 9, but suppressed phosphorylation of STAT3. Epidermal growth factor, a potent stimulator of signal transducer and activator of transcription-3 (STAT3), promoted cell viability and colony formation ability, whereas combination treatment exerted inhibitory effects on epidermal growth factor-induced STAT3 phosphorylation. Finally, HepG2 xenograft mice cotreated with sorafenib and CuB exhibited reduced tumor progression without notable weight loss. In conclusion, sorafenib and CuB exert synergistic antitumor effects through a pathway that may involve STAT3 phosphorylation, and this may represent a promising therapeutic approach for treatment of HCC.

MicroRNA-375 represses tumor angiogenesis and reverses resistance to sorafenib in hepatocarcinoma.

Sorafenib was originally identified as an inhibitor of multiple oncogenic kinases and remains the first-line systemic therapy for advanced hepatocellular carcinoma (HCC). MicroRNAs (miRNAs) have been reported to play critical roles in the initiation, progression, and drug resistance of HCC. In this study, we aimed to identify sorafenib-induced miRNAs and demonstrate their regulatory roles. First, we identified that the expression of the tumor-suppressive miRNA miR-375 was significantly induced in hepatoma cells treated with sorafenib, and miR-375 could exert its antiangiogenic effect partially via platelet-derived growth factor C (PDGFC) inhibition. Then, we demonstrated that sorafenib inhibited PDGFC expression by inducing the expression of miR-375 and a transcription factor, achaete-scute homolog-1 (ASH1), mediated the induction of miR-375 by sorafeinb administration in hepatoma cells. Finally, we verified that the expression of miR-375 was reduced in sorafenib-resistant cells and that the restoration of miR-375 could resensitize sorafenib-resistant cells to sorafenib partially by the degradation of astrocyte elevated gene-1 (AEG-1). In conclusion, our data demonstrate that miR-375 is a critical determinant of HCC angiogenesis and sorafenib tolerance, revealing a novel miRNA-mediated mechanism underlying sorafenib treatment.

C/EBP ß Mediates Endoplasmic Reticulum Stress Regulated Inflammatory Response and Extracellular Matrix Degradation in LPS-Stimulated Human Periodontal Ligament Cells.

Periodontitis is an oral inflammatory disease that not only affects the integrity of local tooth-supporting tissues but also impacts systemic health. A compositional shift in oral microbiota has been considered as the main cause of periodontitis; however, the potential mechanism has not been fully defined. Herein, we investigated the role of CCAAT/enhancer-binding protein ß (C/EBP ß), a member of the C/EBP family of transcription factors, in human periodontal ligament cells (hPDLCs) exposed to Porphyromonas gingivalis (P. gingivalis) lipopolysaccharide (LPS). RT-PCR and Western blotting analysis showed that the expression of C/EBP ß was significantly increased in hPDLCs stimulated with LPS stimuli. Overexpression of C/EBP ß by the recombinant adenoviral vector pAd/C/EBP ß markedly increased the expression of the pro-inflammatory cytokines IL-6 and IL-8, and matrix metalloproteinases (MMP)-8 and -9 in hPDLCs in response to LPS. Furthermore, the activation of endoplasmic reticulum (ER) stress was confirmed in LPS-stimulated hPDLCs by measuring the expression of the ER stress marker molecules protein kinase-like ER kinase (PERK), eIF2α, GRP78/Bip, and C/EBP homologous protein (CHOP). The ER stress inhibitor salubrinal repressed, but inducer tunicamycin enhanced, the production of IL-6, IL-8, MMP-8, and MMP-9 in hPDLCs. Additionally, ER stress inducer tunicamycin significantly increased the expression level of C/EBP ß in hPDLCs. Blocking of C/EBP ß by siRNA resulted in a significant decrease in the secretion of IL-6 and IL-8 and expression of MMP-8 and MMP-9 induced by tunicamycin treatment in hPDLCs. Taken together, ER stress appears to play a regulatory role in the inflammatory response and extracellular matrix (ECM) degradation in hPDLCs in response to LPS stimuli by activating C/EBP ß expression. This enhances our understanding of human periodontitis pathology.

Nicotine induces the production of IL-1ß and IL-8 via the α7 nAChR/NF-κB pathway in human periodontal ligament cells: an in vitro study.

BACKGROUND/AIMS: Tobacco smoking is a major risk factor for the occurrence and progression of periodontitis. We previously demonstrated that nicotine could induce the expression of α7 nicotinic acetylcholine receptors (α7 nAChR) in human and rat periodontal tissues. To further examine the signal pathways mediated by α7 nAChR in periodontal ligament (PDL) cells, we investigated whether nicotine affects interleukin-1ß (IL-1ß) and interleukin-8 (IL-8) via the α7 nAChR/NF-κB pathway in human PDL cells. METHODS: Human PDL cells were pre-incubated with alpha-bungarotoxin (α-BTX) or pyrrolidine dithiocarbamate (PDTC), then cultured with nicotine. Then, we used western blotting, a dual-luciferase reporter, real-time quantitative PCR and an enzyme-linked immunoassay to assess expression of the NF-κB p65 subunit, NF-κB activity and production of IL-1ß and IL-8 in human PDL cells. RESULTS: Compared with the control group, nicotine could significantly induce production of IL-1ß and IL-8 in human PDL cells and cause the similar effects on the expression of the NF-κB p65 subunit and NF-κB activity. CONCLUSION: This study demonstrates that nicotine could induce production of IL-1ß and IL-8 via the α7 nAChR/NF-κB pathway in human PDL cells, providing data for a better understanding of the relationships among smoking, nicotine, and periodontitis.

Oral and craniofacial manifestations and two novel missense mutations of the NTRK1 gene identified in the patient with congenital insensitivity to pain with anhidrosis.

Congenital insensitivity to pain with anhidrosis (CIPA) is a rare inherited disorder of the peripheral nervous system resulting from mutations in neurotrophic tyrosine kinase receptor 1 gene (NTRK1), which encodes the high-affinity nerve growth factor receptor TRKA. Here, we investigated the oral and craniofacial manifestations of a Chinese patient affected by autosomal-recessive CIPA and identified compound heterozygosity in the NTRK1 gene. The affected boy has multisystemic disorder with lack of reaction to pain stimuli accompanied by self-mutilation behavior, the inability to sweat leading to defective thermoregulation, and mental retardation. Oral and craniofacial manifestations included a large number of missing teeth, nasal malformation, submucous cleft palate, severe soft tissue injuries, dental caries and malocclusion. Histopathological evaluation of the skin sample revealed severe peripheral nerve fiber loss as well as mild loss and absent innervation of sweat glands. Ultrastructural and morphometric studies of a shed tooth revealed dental abnormalities, including hypomineralization, dentin hypoplasia, cementogenesis defects and a dysplastic periodontal ligament. Genetic analysis revealed a compound heterozygosity--c.1561T>C and c.2057G>A in the NTRK1 gene. This report extends the spectrum of NTRK1 mutations observed in patients diagnosed with CIPA and provides additional insight for clinical and molecular diagnosis.

Conservation of root-fractured primary teeth--report of a case.

A 3.5-year-old girl presented to our clinic experiencing pain in her maxillary central incisors following traumatic injury during a fall. Radiographic examination revealed both primary maxillary central incisors with mid-root and apical third horizontal root fractures, respectively. Splinting with orthodontic brackets and stainless steel wire was performed. At 2 weeks, resorption of the apical fragments in both injured teeth was observed, and after 3 months, almost complete resorption was noted on radiographs. Tooth mobility at this point was back to physiologic levels and the splint was removed. After 2.5 years, the primary maxillary incisors were replaced by permanent incisors demonstrating normal tooth color, position, and root development. Although this case illustrated the favorable prognosis of two primary teeth with root fractures and severely mobile coronal fragments by a conservative approach, more scientific evidences are needed and frequent recalls are necessary when primary root fractures are attempted to be managed with splinting.

Cementum- and periodontal ligament-like tissue formation by dental follicle cell sheets co-cultured with Hertwig's epithelial root sheath cells.

Dental follicle cells (DFCs) are believed contain the precursor cells of the periodontium and can form cell sheets by secreting extracellular matrix (ECM) proteins. Cell sheet engineering has been recently developed and applied successfully in the field of tissue regeneration. However, research on the in vitro characteristics of DFC sheets is lacking and an assessment of whether DFC sheets can produce periodontal tissues in vivo has not been reported. To test the characteristics and applicability of DFC sheets in this field, we established a co-culture system of rat DFCs and Hertwig's epithelial root sheath (HERS) cells in vitro, and included the following controls: a co-culture of DFCs and alveolar mucosa epithelial cells, DFCs with no cells in the upper chamber, and DFCs cultured without an upper chamber. After 3 weeks of co-culturing the cells, the DFC sheets were transplanted into adult male rats' omenta. One week after co-culturing DFCs with HERS cells, mRNA levels of collagen type I (COL-1), alkaline phosphatase (ALP), runt related transcription factor 2 (Runx 2) and bone sialoprotein (BSP) were increased significantly. In addition, after 3 weeks of co-culturing the cells, the numbers of ALP-, osteocalcin (OCN)-, BSP- and osteoprotegerin (OPG)-positive DFCs increased. The DFCs also produced more calcified nodules and exhibited an increased number of subcellular organelles, which are important for protein synthesis and secretion. Moreover, gap junctions were found between the experimental DFCs within the sheet. Five weeks of in vivo growth of DFC sheets pre-exposed to HERS cells led to the formation of cementum-like tissues, which were positive for OCN, BSP and OPG, as well as the formation of periodontal ligament-like tissues, which were positive for COL-1. In contrast, control cells only produced fibrous tissues. These results indicate that the DFC sheets induced by HERS cells are able to produce periodontal tissues through epithelial-mesenchymal interactions. Therefore, DFC sheets may be useful in the field of periodontium regeneration.

Formation of bone-like tissue by dental follicle cells co-cultured with dental papilla cells.

During tooth root formation, dental follicle cells (DFCs) differentiate into osteoblasts/cementoblasts when they are in contact with pre-existing dentin. Since some factors of dentin matrix were also produced by dental papilla cells (DPCs) and could induce DFCs differentiation, we hypothesized that DPCs can directly promote DFCs differentiation and that differentiation could occur in a co-culture model. To test this hypothesis, we investigated the characteristics of DFCs that are influenced by DPCs in an in vitro co-culture and in vivo heterotopic transplant model. One week into the co-culture, there were significant increases in the mRNA level of bone morphogenetic protein 2 (BMP2), osteoprotegerin (OPG), bone sialoprotein (BSP) and osteocalcin (OCN), and a decrease of the receptor activator of nuclear factor κB ligand (RANKL). Additionally, the number of BMP2-, OPG-, BSP- and OCN-positive DFCs increased whereas RANKL-positive DFCs decreased. Three weeks after co-culture, DFCs produced calcified nodules, accompanied with increased sub-cellular organelles for protein synthesis and secretion. In the heterotopic transplant model, the adult male rats were used as hosts, DFCs were transplanted into the omentum. In vivo 5-week growth of DFCs in the presence of DPCs led to the formation of bone-like tissues, positive for BSP, OCN and BMP2. In contrast, DFCs alone led to fibrous-like tissues. These results indicated that in the absence of pre-existing dentin, DPCs can stimulate osteogenesis and inhibit osteoclastogenesis in DFCs and suggested a novel strategy to promote DFCs differentiation.

Inhibition of RANK/RANKL signal transduction pathway: a promising approach for osteoporosis treatment.

Osteoporosis is a bone disease causing impaired bone strength. It is characterized by increased osteoclast formation or enhanced bone resorption, leading to an increased risk of fragility fractures. Its prevalence increases with age. The advent of an aging population suggests that progressively more individuals will develop this disease in the aging population. A number of drugs for the prevention and treatment of osteoporosis act by inhibiting bone resorption. However, the effectiveness of osteoporosis treatment in clinical practice is limited. Since the osteoclast is the only cell in the body that is capable of resorbing bone, understanding its biology will be necessary for developing a new therapeutic approach for osteoporosis. Recently, it was discovered that the receptor activator of nuclear factor kappaB (RANK)/RANK ligand (RANKL)/osteoprotegerin (OPG) system is an important signal transduction pathway that regulates osteoclast formation. The binding of OPG to RANKL inhibits the binding between RANKL and RANK; this, in turn, prevents osteoclast precursors from differentiating and fusing to form mature osteoclasts. Therefore, the inhibition of the RANK/RANKL pathway inhibits osteoclast formation, differentiation, activation, and bone resorption. A potential clinical antiresorptive therapy can be developed by using an anti-RANKL monoclonal antibody, such as denosumab, that binds to RANKL with high affinity and specificity and blocks RANKL-RANK interactions.