Self-Regenerating Photothermal Agents for Tandem Photothermal and Thermodynamic Tumor Therapy.

Small molecule-based photothermal agents (PTAs) hold promising future for photothermal therapy; however, unexpected inactivation exerts negative impacts on their application clinically. Herein, a self-regenerating PTA strategy is proposed by integrating 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) radical cation (ABTS•+) with a thermodynamic agent (TDA) 2,2'-azobis[2-(2-imidazolin-2-yl) propane] dihydrochloride (AIPH). Under NIR laser, the photothermal effect of ABTS•+ accelerates the production of alkyl radicals by AIPH, which activates the regeneration of ABTS•+, thus creating a continuous positive feedback loop between photothermal and thermodynamic effects. The combination of ABTS•+ regeneration and alkyl radical production leads to the tandem photothermal and thermodynamic tumor therapy. In vitro and in vivo experiments confirm that the synergistic action of thermal ablation, radical damage, and oxidative stress effectively realizes tumor suppression. This work offers a promising approach to address the unwanted inactivation of PTAs and provides valuable insights for optimizing combination therapy.

Multiorgan dysfunction syndrome due to high-dose cantharidin poisoning: A case report.

BACKGROUND: This report delves into the diagnostic and therapeutic journey undertaken by a patient with high-dose cantharidin poisoning and multiorgan dysfunction syndrome (MODS). Particular emphasis is placed on the comprehensive elucidation of the clinical manifestations of high-dose cantharidin poisoning, the intricate path to diagnosis, and the exploration of potential underlying mechanisms. CASE SUMMARY: A patient taking 10 g of cantharidin powder orally subsequently developed MODS. The patient was treated with supportive care, fluid hydration and antibiotics, and hemoperfusion and hemofiltration therapy for 24 h and successfully recovered 8 d after hospital admission. Cantharidin poisoning can cause life-threatening MODS and is rare clinically. This case underscores the challenge in diagnosis and highlights the need for early clinical differentiation to facilitate accurate assessment and prompt intervention. CONCLUSION: This article has reported and analyzed the clinical data, diagnosis, treatment, and prognosis of a case of high-dose cantharidin poisoning resulting in MODS and reviewed the relevant literature to improve the clinical understanding of this rare condition.

Hypocapnia is an independent predictor of in-hospital mortality in acute heart failure.

AIMS: Acute heart failure (AHF) poses a major threat to hospitalized patients for its high mortality rate and serious complications. The aim of this study is to determine whether hypocapnia [defined as the partial pressure of arterial carbon dioxide (PaCO2 ) below 35 mmHg] on admission could be associated with in-hospital all-cause mortality in AHF. METHODS AND RESULTS: A total of 676 patients treated in the coronary care unit for AHF were retrospectively analysed, and the study endpoint was in-hospital all-cause mortality. The 1:1 propensity score matching (PSM) analysis, Kaplan-Meier curve, and Cox regression model were used to explore the association between hypocapnia and in-hospital all-cause mortality in AHF. Receiver operating characteristic (ROC) curve and Delong's test were used to assess the performance of hypocapnia in predicting in-hospital all-cause mortality in AHF. The study cohort included 464 (68.6%) males and 212 (31.4%) females, and the median age was 66 years (interquartile range 56-74 years). Ninety-eight (14.5%) patients died during hospitalization and presented more hypocapnia than survivors (76.5% vs. 45.5%, P < 0.001). A 1:1 PSM was performed between hypocapnic and non-hypocapnic patients, with 264 individuals in each of the two groups after matching. Compared with non-hypocapnic patients, in-hospital mortality was significantly higher in hypocapnic patients both before (22.2% vs. 6.8%, P < 0.001) and after (20.8% vs. 8.7%, P < 0.001) PSM. Kaplan-Meier curve showed a significantly higher probability of in-hospital death in patients with hypocapnia before and after PSM (both P < 0.001 for the log-rank test). Multivariate Cox regression analysis showed that hypocapnia was an independent predictor of AHF mortality both before [hazard ratio (HR) 2.22; 95% confidence interval (CI) 1.23-3.98; P = 0.008] and after (HR 2.19; 95% CI 1.18-4.07; P = 0.013) PSM. Delong's test showed that the area under the ROC curve was improved after adding hypocapnia into the model (0.872, 95% CI 0.839-0.901 vs. 0.855, 95% CI 0.820-0.886, P = 0.028). PaCO2 was correlated with the estimated glomerular filtration rate (r = 0.20, P = 0.001), left ventricular ejection fraction (r = 0.13, P < 0.001), B-type natriuretic peptide (r = -0.28, P < 0.001), and lactate (r = -0.15, P < 0.001). Kaplan-Meier curve of PaCO2 tertiles and multivariate Cox regression analysis showed that the lowest PaCO2 tertile was associated with increased risk of in-hospital mortality in AHF (all P < 0.05). CONCLUSIONS: Hypocapnia is an independent predictor of in-hospital mortality for AHF.

A Cohort Study on Deficiency of ADA2 from China.

PURPOSE: Deficiency of adenosine deaminase 2 (DADA2), an autosomal recessive autoinflammatory disorder caused by biallelic loss-of-function variants in adenosine deaminase 2 (ADA2), has not been systemically investigated in Chinese population yet. We aim to further characterize DADA2 cases in China. METHODS: A retrospective analysis of patients with DADA2 identified through whole exome sequencing (WES) at seventeen rheumatology centers across China was conducted. Clinical characteristics, laboratory findings, genotype, and treatment response were analyzed. RESULTS: Thirty patients with DADA2 were enrolled between January 2015 and December 2021. Adenosine deaminase 2 enzymatic activity was low in all tested cases to confirm pathogenicity. Median age of disease presentation was 4.3 years and the median age at diagnosis was 7.8 years. All but one patient presented during childhood and two subjects died from complications of their disease. The patients most commonly presented with systemic inflammation (92.9%), vasculitis (86.7%), and hypogammaglobinemia (73.3%) while one patient presented with bone marrow failure (BMF) with variable cytopenia. Twenty-three (76.7%) patients were treated with TNF inhibitors (TNFi), while two (6.7%) underwent hematopoietic stem cell transplantation (HSCT). They all achieved clinical remission. A total of thirty-nine ADA2 causative variants were identified, six of which were novel. CONCLUSION: To establish early diagnosis and improve clinical outcomes, genetic screening and/or testing of ADA2 enzymatic activity should be performed in patients with suspected clinical features. TNFi is considered as first line treatment for those with vascular phenotypes. HSCT may be beneficial for those with hematological disease or in those who are refractory to TNFi.

Association between Serum Oxytocin, Bone Mineral Density and Body Composition in Chinese Adult Females.

Background and Objectives: Oxytocin (OT) is a neuropeptide hormone which is known for its classical effects in pregnancy and lactation. Recently, growing evidence demonstrated a close relation between OT and bone. The present study aimed to explore the relationship between OT, bone and osteoporosis risk in Chinese adult females. Materials and Methods: in total, 149 adult females were enrolled. The serum OT levels were measured using ELISA kits. Bone mineral density (BMD) and body composition were measured by dual-energy X-ray absorptiometry (DXA). The study subjects were divided into two groups according to their menopause status and then divided into tertiles based on their serum OT level. Results: Serum OT, serum estradiol and BMD at three skeletal sites were significantly higher in the premenopausal group than in the postmenopausal group (p < 0.001, p = 0.008 and p < 0.001, respectively). In the tertile analysis, relative to tertile 1, significant associations were found for tertile 3 for OT levels and higher BMD in the femoral neck and total hip, in both pre- and postmenopausal groups. Using logistic regression analysis, tertile 3 appeared less likely to have low-BMD osteoporosis than tertile 1 (OR = 0.257, 95% CI = 0.073, 0.910). In multivariate stepwise regression analysis, OT and total lean mass were two positive determinants of BMD in the femoral neck and total hip in the premenopausal group (adjusted R2 for the model = 0.232 and 0.199, respectively; both p < 0.001). Conclusion: Our study demonstrated positive associations between serum OT levels and BMD in a Chinese (non-Caucasian) population. OT appeared to be more strongly associated with hip BMD in premenopausal females. These results may suggest a protective role and potential therapeutic use of OT in osteoporosis, especially for premenopausal women.

Lipid droplet-hitchhiking probe creates Trojan foam cells for fluorescence/photoacoustic imaging of atherosclerotic plaques.

Since atherosclerosis, a disease characterized by abnormal arterial lipid deposition, may lead to fatal cardiovascular diseases, imaging of atherosclerotic plaques is of great value for their pathological assessment. In this study, we propose a lipid droplet (LD)-hitchhiking strategy to in situ create Trojan foam cells for fluorescence/photoacoustic imaging of atherosclerotic plaques via homologous targeting effect. In our design, functional liposomes (DCP liposomes) composed of phospholipid dioleoylphosphatidylserine (DOPS), a novel LD inducer we found, and Cypate-PC, a synthesized lipid-like molecular probe, have demonstrated great capability of inducing LDs in monocytes/macrophages while being enveloped into the resulting Trojan foam cells. Taking advantage of homologous targeting effect, the imaging probe hitchhikes on the LDs in Trojan foam cells for targeted transport to the plaque sites. Moreover, the confinement in highly hydrophobic LDs endows the imaging probe with high efficiency in light absorption, enabling greatly intensified fluorescence/photoacoustic signals. The DCP liposomes have shown great potency in inducing the generation of Trojan foam cells, and eventually ex vivo fluorescence imaging and in vivo photoacoustic imaging of atherosclerotic plaques. The proposed strategy provides more insights into the design of targeted imaging methodologies, and also an effective avenue to facilitate the evaluation and subsequent treatment of atherosclerotic plaques.

Astragaloside IV inhibits protein tyrosine phosphatase 1B and improves insulin resistance in insulin-resistant HepG2 cells and triglyceride accumulation in oleic acid (OA)-treated HepG2 cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Astragaloside IV (AST IV) is the active component of Astragalus membranaceus (Fisch.) Bunge, which regulates lipid and carbohydrate metabolism and improves insulin resistance. In this study, we investigated the effects of AST IV on insulin resistant cells and a non-alcoholic fatty liver disease (NAFLD) model induced by high-concentration insulin or oleic acid (OA) in HepG2 cells, as well as the associated regulatory markers. METHODS: First, the target of AST IV was predicted via pharmacophore model matching and molecular docking. Then, enzyme kinetics experiments were conducted in vitro to determine the effect of AST IV on the target protein. Next, AST IV's toxicity was tested on HepG2 cells in vitro, through an insulin resistance model and an NAFLD model, by high-concentration insulin or OA, respectively. To explore the effects of AST IV on insulin resistance and lipid metabolism, we detected the related indexes of glucose and lipid metabolism through commercially available kits. Relevant proteins were also detected by Western blot to provide future direction for study. RESULTS: Our preliminary results of pharmacophore model matching and molecular docking suggested that AST IV and protein tyrosine phosphatase 1B (PTP1B) can be well-combined through hydrogen bonding. Further, the enzyme kinetics experiment showed that AST IV was an effective and specific inhibitor to PTP1B. We found that the protein level of PTP1B in HepG2 cells was significantly increased after treating with high-concentration insulin or OA. Additionally, the intervention of AST IV significantly increased glucose consumption in an insulin resistance model and reduced the content of triglyceride (TG), total cholesterol (TC), and free fatty acid (FFA) in the NAFLD model. Moreover, the 2-N-(7-nitrobenze-2-oxa-1, 3 diazol-4-yl) (2-NBDG) uptake rate in the NAFLD model was also greatly improved. These results validated the effects of AST IV on improving insulin resistance and lipid accumulation. Furthermore, Western blot results illustrated that AST IV suppressed PTP1B and increased levels of phosphorylated insulin receptor (p-IR) and phosphorylated insulin receptor substrate-1 (p-IRS-1) in insulin-resistant HepG2 cells, while also decreasing protein levels of PTP1B and sterol element regulatory binding protein-1c (SREBP-1c) in the NAFLD model. CONCLUSION: This study demonstrated that AST IV inhibited PTP1B and effectively improved insulin resistance in insulin-resistant HepG2 cells and triglyceride accumulation in OA-treated HepG2 cells.

[A Meta-analysis of the effect of non-surgical periodontal therapy on inflammatory factors in patients with chronic kidney disease and periodontitis].

OBJECTIVE: A study was conducted to systematically evaluate the clinical efficacy of inflammatory factors in patients with chronic kidney disease and periodontitis after non-surgical periodontal therapy. METHODS: We searched the databases of CNKI, Wanfang, CBM, PubMed, Embase, and Cochrane Library from inception to December 2019. Two reviewers independently collected all literature related to inflammatory factors in patients with chronic kidney disease and periodontitis after non-surgical periodontal therapy. These factors include C-reactive protein (CRP), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α). The literature was screened according to the inclusion and exclusion criteria. The quality of the studies was strictly evaluated, and the data were extracted. The literature of randomized controlled trials in accordance with the standards was Meta-analyzed with Revman 5.3 software. RESULTS: Six randomized controlled trials were included. Compared with the control groups, the results of meta-analysis showed that non-surgical periodontal therapy significantly reduced the levels of CRP [MD=-0.58, 95%CI (-1.13, -0.02), P=0.04] and IL-6 [MD=-2.76, 95%CI (-5.15, -0.37), P=0.02] in these patients but not that of TNF-α [MD=-3.87, 95%CI (-8.79, 1.05), P=0.12]. CONCLUSIONS: Simultaneous regular renal treatment and non-surgical periodontal therapy can help relieve the periodontal damage on patients with chronic kidney disease and periodontitis. Moreover, it can improve the status of some inflammatory factors. This finding is conducive to the control and treatment of chronic kidney disease and periodontitis and needs to be a focus of research and in clinical operation.

[Effect of acupoint catgut embedding on levels of PG-related factors and NF-κB proteins in ute-rine tissues of rats with primary dysmenorrhea].

OBJECTIVE: To observe the effect of acupoint catgut embedding on levels of prostaglandin F2α(PGF2α), cyclooxygenase-2(COX-2) and nuclear factor kappa B(NF-κB) in the uteruses of rats with primary dysmenorrhea (PD), so as to explore its mechanisms underlying improvement of PD. METHODS: Forty female SD rats were randomly divided into normal, model, acupoint catgut embedment and medication groups, with 10 rats in each group. The PD model was established by subcutaneous injection of estradiol benzoate (0.5 mg on the 1st day, and 0.2 mg thereafter) once daily for 10 days, followed by i．p. of oxytocin 2 U(0.5 mg•5 U－1•mL－1) on the 11th day. Catgut embedment was applied to "Guanyuan"(CV4) and "Sanyinjiao"(SP6) on day 1 and 5 while modeling, and rats of the medication group received gavage of ibuprofen (1.25 mg/mL, 0.8 mL/rat) once daily for 10 d. The level of PGF2αin the uterus tissuewas assayed by ELISA, and the expression levels of uterine COX-2, phospho (p)-NF-κB p65, NF-κB p65 proteins were detected by Western blot. RESULTS: Compared with the normal group, the levels of PGF2α， COX-2, p-NF-κB p65 and NF-κB p65 in the uterine tissues were significantly increased in the model group (P<0.05). Compared with the model group, the levels of PGF2α and COX-2 in both catgut embedment and medication groups as well as p-NF-κB p65 in the catgut embedment group were significantly decreased (P<0.05). Compared with the medication group, the level of p-NF-κB 65 in the catgut embedment group was significantly decreased (P<0.05). CONCLUSION: The analgesic effect of acupoint catgut embedding may be related to its effects in down-regulating the expression of p-NF-κB and the levels of COX-2 and PGF2αin uteruses of PD rats.

[Astragaloside IV attenuates cerebral ischemia and reperfusion injury and reduces activation of NLRP3 inflammasome and NF-κB phosphorylation in rats following a transient middle cerebral artery occlusion].

The present study was aimed to investigate the protective effect and anti-inflammation mechanism of astragaloside IV (AST-IV) on cerebral ischemia and reperfusion injury. Following the establishment of cerebral ischemia and reperfusion model in rats by modified suture method, neurological deficit scores and cerebral infarct volume were used to evaluate the pharmacological effect of AST-IV against cerebral ischemia-reperfusion injury. Western blot was used to detect the expression levels of NLRP3, pro-Caspase-1, Caspase-1, pro-IL-1ß, IL-1ß, pro-IL-18, IL-18, phosphorylated and total nuclear factor kappa B (NF-κB)/p65 protein in the brain tissue. The results showed that compared with model group, the intervention of AST-IV decreased the neurological deficit scores, reduced the cerebral infarct volume, decreased the levels of NLRP3, Caspase-1, pro-IL-1ß, IL-1ß, pro-IL-18 and IL-18, and inhibited the expression of phosphorylated NF-κB in brain tissue. The results suggest that AST-IV has a protective effect against cerebral ischemia and reperfusion injury, and its mechanism is related to inhibiting the phosphorylation of NF-κB and NLRP3 inflammasome activation.

Cornel Iridoid Glycoside Improves Locomotor Impairment and Decreases Spinal Cord Damage in Rats.

Purpose. This study was to investigate the effects of cornel iridoid glycoside (CIG) on spinal cord injury (SCI) in rats. Methods. The thoracic cord (at T9) of rats was injured by clip compression for 30 sec. Locomotor function was assessed using the Basso, Beattie, and Bresnahan (BBB) rating scale. Neuroanatomic stereological parameters as well as Nogo-A, p75 neurotrophin receptor (p75NTR), and ROCKII expression were measured by histological processing, immunohistochemistry, and stereological analyses. The axons passing through the lesion site were detected by BDA tracing. Results. Intragastric administration of CIG (60 and 180 mg/kg) improved the locomotor impairment at 10, 17, 24, and 31 days post-injury (dpi) compared with untreated SCI model rats. CIG treatment decreased the volume of the lesion epicenter (LEp) and increased the volume of spared tissue and the number of surviving neurons in the injured spinal cord at 31 dpi. CIG promoted the growth of BDA-positive axons and their passage through the lesion site and decreased the expression of Nogo-A, p75NTR, and ROCKII both in and around the LEp. Conclusion. CIG improved the locomotor impairment, decreased tissue damage, and downregulated the myelin-associated inhibition signaling pathway in SCI rats. The results suggest that CIG may be beneficial for SCI therapy.

Imaging features of primary hepatic leiomyosarcoma: A case report and review of literature.

Primary hepatic leiomyosarcoma (PHL) is an extremely rare tumour. This tumour is difficult to diagnose by imaging examinations due to its rarity, and non-specific conventional imaging manifestations and clinical presentation. The present study reports the case of a 42-year-old male with PHL that was confirmed by histopathological and immunohistochemical examinations. Multimodal imaging examinations, including ultrasound, computed tomography (CT), magnetic resonance imaging (MRI), positron emission tomography-CT and digital subtraction angiography, were performed. The imaging manifestations were analysed and the associated literature was reviewed. The results found that no characteristic imaging appearance was present on ultrasound or plain CT scan. However, on unenhanced MRI, the tumours presented with a heterogeneous low signal density on T1-weighted imaging (WI) and a high signal density on T2WI and diffusion-WI. On gadopentetate dimeglumine enhanced MRI, the lesions were not enhanced during the arterial and portal venous phases; by contrast, these lesions were evidently enhanced during the 5-min delayed phase. Therefore, the delayed imaging of enhanced MRI is likely to be used to differentiate PHL from other hepatic tumours.

Clinical, molecular, and T cell subset analyses in a small cohort of Chinese patients with hyper-IgM syndrome type 1.

Type 1 hyper-IgM syndrome (HIGM1) is a rare primary immunodeficiency disease caused by mutations in the CD40L gene. Patients often present with recurrent infections and autoimmune manifestations. We investigated the clinical and molecular characteristics of HIGM1 in thirteen patients from the Chinese mainland and examined the proportion of CD4(+)CD25(+)FoxP3(+)Treg, Th17, and Th1 cells in the peripheral blood. We identified ten distinct CD40L mutations in eleven patients: one missense mutation, one nonsense mutation, one insertion mutation (in frame), and seven deletions. Six of these mutations were novel. We observed the percentage of Tregs in the peripheral blood of HIGM1 patients decreased markedly compared with that in healthy controls, but no statistically significant differences was found in the percentages of Th17 and Th1. The identified mutations reflect the heterogeneity of the CD40L gene in HIGM1. Precise genetic diagnosis of HIGM1 will enable appropriate therapeutic interventions, reliable detection of carriers, and genetic counseling. Skewed Treg, Th17/Treg, and Th1/Treg profiles may be associated with immune responses to autoimmunity or infection, which requires replication in larger studies.

Superior laryngeal nerve loop: patterns and surgical implications.

This study clarifies the patterns of the superior laryngeal nerve loop (SLN loop), connecting the cervical sympathetic chain (CSC) and the SLN and its branches, so as to provide an anatomic basis for decreasing the risk of injury to the external laryngeal nerve (ELN) during neck surgery. Fifty Chinese adult human cadavers fixed with 4 % formalin were dissected, and their SLN loop patterns were analyzed and summarized. In 98 of 100 sides the CSC anastomosed with the SLN and its branches, forming a looped nerve structure which we called the SLN loop. The SLN loops could be divided into five types: e ( n ), t ( n ), i ( n ), t ( n ) e ( n ), and i ( n ) e ( n ) based on morphological variations. The results demonstrated that e ( n ) was most frequently found in the samples (82/100) followed by t ( n ) (9/100), i ( n ) (3/100), t ( n ) e ( n ) (2/100), and i ( n ) e ( n ) (2/100). Comparing with the previous work, we identified additional 18 subtypes of the SLN loop. The relations of the SLN loop to the surrounding structures were complicated, which brought more challenges to thyroidectomy. Thus, we do not advocate routine identification of ELN/ELN loop during the process of thyroidectomy, especially systematic identification of ELN during operation. However, this study introduces the possibility that nerve injury can be avoided by exposure of the nerve via careful dissection in the region of the superior pole of the thyroid gland to the extent that we can initiate individual ligation of the superior polar vessels, along with the help of neuromonitors, video monitors, and magnifying loupes.

An applied anatomical study on the recurrent laryngeal nerve and inferior thyroid artery.

PURPOSE: The aim of this study was to provide some important information about the morphology and topography of the recurrent laryngeal nerve (RLN) and inferior thyroid artery (ITA), which significantly helps localize and protect the RLN in neck surgery, especially in thyroid surgery. METHODS: Eighty adult cadavers (160 sides) fixed with formalin were dissected, analyzed and measured. RESULTS: (1) 87.5% of the RLNs gave off multiple branches like a tree; the incidence of the RLN loop, connecting one branch to another was 3.125%; in 9.375%, one branch of RLN combined with cervical sympathetic chain (CSC) or superior laryngeal nerve (SLN). (2) A double RLN appeared in four sides, a non-recurrent inferior laryngeal nerve appeared in two cases. (3) In two cases, the RLN communicated with both of the SLN and the CSC near thyroid gland. (4) Most of the ITAs was derived from thyrocervical trunk, and divided into two or three branches before entering the thyroid gland. (5) Three ITAs gave off esophageal branch, one ITA gave off tracheal branch, one right ITA originated abnormally. (6) On the left side, the RLN was behind the ITA in 86.25% of the cases, in front of the artery in 7.5%, the nerve was between artery branches in 2.5%, the artery was between nerve branches in 1.25%, and was among the combined in 2.5%. On the right side, the RLN was in front of the artery in 75.0%, behind the artery in 10.0%, among the branches of the artery in 5.0%, 10.0% the branches of both nerves and artery were interlaced that the relationship between the branches of the nerve and the artery was uncertain. CONCLUSIONS: Because of the variability of the RLN and ITA and the complicated relationship between them, it is necessary to dissect and recognize the RLN to avoid mistaking, ignoring, and misligating of the nerve before ligating the ITA.

Autocrine/paracrine action of vitamin D on FGF23 expression in cultured rat osteoblasts.

To explore the local mechanisms of fibroblast growth factor (FGF) 23 regulations, we examined the FGF23 expression patterns in an osteoblast culture model. The characteristics of cultured rat calvaria osteoblasts in half-confluence, confluence, osteoid deposition, and osteoid mineralization stages might reflect the proliferation, differentiation, maturation, and matrix mineralization stages, respectively. Compared with proliferating cells in half-confluence, FGF23 expression was upregulated by 7.5-fold at the mRNA level and 126% at the protein level in confluent differentiated cells as determined by real-time RT-PCR and Western blot analysis. Interestingly, mRNA levels of CYP27B1 (the gene coding for 1alpha-hydroxylase enzyme which catalyses the conversion of 1alpha,25-dihydroxyvitamin D, 1alpha,25[OH]2D, from its inactive form, 25-hydroxycholecalciferol, 25[OH]D) and CYP24A (the gene coding for 24-hydroxylase, a target gene of 1alpha,25[OH]2D) were significantly increased by twofold and 34-fold, respectively, in differentiated osteoblasts compared with proliferating cells. We next examined if the local production of 1alpha,25(OH)2D might contribute to the FGF23 upregulation. We cultured osteoblasts in serum-free medium with or without 25-(OH)D (the substrate of 1alpha-hydroxylase). FGF23 mRNA levels were increased in proliferating cells (16-fold) and in differentiated cells (28-fold) by the physiological dose of 25-(OH)D3 treatment. CYP27B1 was slightly but significantly upregulated and CYP24A was increased by 1,700-fold and 800-fold, respectively, in transcriptional levels. Because FGF23 was upregulated in confluent osteoblasts regardless of the presence or absence of 25-(OH)D in serum-free medium, we further examined the possible impact of cell communication on FGF23 expression. We treated osteoblasts with carbenoxolone, a gap junction Cx43 blocker in serum-free medium. The FGF23 mRNA level was reduced by 50% in confluent differentiated cells and slightly but not significantly reduced in half-confluent cells by carbenoxolone treatments. The results suggested that upregulation of FGF23 in differentiated osteoblast appeared to be due to increased autocrine/paracrine action of osteoblast-derived 1alpha,25(OH)2D and increased cell communication, which were identified in cultured rat calvaria osteoblasts. These results indicate that FGF23 expression was stimulated not only by circulating calcitriol but also by locally produced 1alpha,25(OH)2D. The local mechanisms of FGF23 expression remain to be characterized.