Involvement of Macrophages and Spinal Microglia in Osteoarthritis Pain.

PURPOSE OF REVIEW: Chronic pain in osteoarthritis (OA) is characterized by pain sensitization, which involves both peripheral and central mechanisms. Studies suggest synovial macrophage and spinal microglia are implicated in pain sensitization in OA. We, therefore, reviewed the evidence of whether synovial macrophage and spinal microglia facilitated pain sensitization at diverse levels and how this event occurred in OA. RECENT FINDINGS: Peripherally, joint inflammation is now believed to be a source of OA-related pain. Synovial macrophages accumulate in OA inflamed synovium and display a pro-inflammatory phenotype. Abundant macrophage-derived pro-inflammatory cytokines and other pain-causing substance facilitate hyperexcitation of primary sensory neuron in OA-related pain. Thus, activated synovial macrophage was considered a predictor for phenotyping of OA pain clinically. In response to affected joint-derived strong nociception, aberrant neuronal excitability is often associated with the hyperactivity of microglia in the spinal dorsal horn, thereby leading to central sensitization. Hyperactivity of synovial macrophage and spinal microglia underlies the mechanisms of pain sensitization at the peripheral and central level in OA. This concept provides not only a clinically relevant strategy for identifying the phenotype of OA-related pain but also has the potential to develop individualized interventions for OA, particularly in those patients with hyperactivity of macrophage and microglia.

Posttranslational lysine 2-hydroxyisobutyrylation of human sperm tail proteins affects motility.

STUDY QUESTION: Does lysine 2-hydroxyisobutyrylation, a newly identified protein posttranslational modification (PTM), occur in human sperm and affect human sperm function? SUMMARY ANSWER: Lysine 2-hydroxyisobutyrylation mainly occurs in human sperm tail proteins, and excessive lysine 2-hydroxyisobutyrylation affects human sperm motility. WHAT IS KNOWN ALREADY: PTM is regarded as an important pathway in regulating sperm function since mature sperm are almost transcriptionally silent. However, only phosphorylation was extensively studied in mature sperm to date. Lysine 2-hydroxyisobutyrylation, a newly characterised PTM, is broadly conserved in both eukaryotic and prokaryotic cells. Although histone lysine 2-hydroxyisobutyrylation has been shown to be associated with active gene expression in spermatogenic cells, the presence, regulatory elements and function of lysine 2-hydroxyisobutyrylation have not been characterised in mature sperm. STUDY DESIGN, SIZE, DURATION: Sperm samples were obtained from normozoospermic men and asthenozoospermic men who visited the reproductive medical centre at Jiangxi Provincial Maternal and Child Health Hospital, Nanchang, Jiangxi, China, between May 2017 and November 2018. In total, 58 normozoospermic men and 65 asthenozoospermic men were recruited to participate in this study. PARTICIPANTS/MATERIALS, SETTING, METHODS: Lysine 2-hydroxyisobutyrylation was examined using immunoblotting and immunofluorescence assays using a previously qualified pan anti-lysine 2-hydroxyisobutyrylation antibody. The immunofluorescence assay was imaged using super-resolution structured illumination microscopy. Sperm viability was examined by using the eosin staining method, and sperm motility parameters were assessed by computer-assisted sperm analysis. Sperm penetration ability was determined by evaluating the ability of the sperm to penetrate a 1% (w/v) methylcellulose solution. The level of intracellular adenosine triphosphate (ATP) was detected using a rapid bioluminescent ATP assay kit. MAIN RESULTS AND THE ROLE OF CHANCE: Lysine 2-hydroxyisobutyrylation was present in several proteins (20-100 kDa) mainly located in the tail of human sperm. Sperm lysine 2-hydroxyisobutyrylation was derived from 2-hydroxyisobutyrate (2-Hib) and was regulated by acyltransferase P300 and nicotinamide adenine dinucleotide-dependent lysine deacylase sirtuins. Elevation of sperm lysine 2-hydroxyisobutyrylation by 2-Hib decreased total motility, progressive motility, penetration ability and ATP level of human sperm. Interestingly, the level of sperm lysine 2-hydroxyisobutyrylation was higher in asthenozoospermic men than that in normozoospermic men and was negatively correlated with the progressive motility of human sperm. Furthermore, high levels of lysine 2-hydroxyisobutyrylation in asthenozoospermic men accompanied decreased ATP levels. LIMITATIONS, REASONS FOR CAUTION: Although the present study indicated the involvement of sperm lysine 2-hydroxyisobutyrylation in regulating human sperm motility, the underlying mechanism needs to be further illustrated. WIDER IMPLICATIONS OF THE FINDINGS: The findings of this study provide insight into the novel role of lysine 2-hydroxyisobutyrylation in human sperm and suggest that abnormality of sperm lysine 2-hydroxyisobutyrylation may be one of the causes for asthenozoospermia. STUDY FUNDING/COMPETING INTEREST(S): National Natural Science Foundation of China (81771644 to T.L. and 81871207 to H.C.); Natural Science Foundation of Jiangxi province (20171ACB21006). The authors have no conflicts of interest to declare.

Lysine glutarylation in human sperm is associated with progressive motility.

STUDY QUESTION: Is there a role for lysine glutarylation (Kglu), a newly identified protein post-translational modification (PTM), in human sperm? SUMMARY ANSWER: Kglu occurs in several proteins located in the tail of human sperm, and it was reduced in asthenozoospermic (A) men and positively correlated with progressive motility of human sperm, indicating its important role in maintaining sperm motility. WHAT IS KNOWN ALREADY: Since mature sperm are almost transcriptionally silent, PTM is regarded as an important pathway in regulating sperm function. However, only phosphorylation has been extensively studied in mature sperm to date. Protein lysine modification (PLM), a hot spot of PTMs, was rarely studied except for a few reports on lysine methylation and acetylation. As a newly identified PLM, Kglu has not been well characterized, especially in mature sperm. STUDY DESIGN, SIZE, DURATION: Sperm samples were obtained from normozoospermic (N) men and A men who visited the reproductive medical center between February 2016 and January 2018. In total, 61 N men and 59 A men were recruited to participate in the study. PARTICIPANTS/MATERIALS, SETTING, METHODS: Kglu was examined by immunoblotting and immunofluorescence assays using a previously qualified pan-anti-glutaryllysine antibody that recognizes glutaryllysine in a wide range of sequence contexts (both in histones and non-histone substrates) but not the structurally similar malonyllysine and succinyllysine. The immunofluorescence assay was imaged using laser scanning confocal microscopy and super-resolution structured illumination microscopy. Sperm motility parameters were examined by computer-assisted sperm analysis. MAIN RESULTS AND THE ROLE OF CHANCE: Kglu occurs in several proteins (20-150 kDa) located in the tail of human sperm, especially in the middle piece and the latter part of the principal piece. Sperm Kglu was modulated by regulatory systems (enzymes and glutaryl-CoA) similar to those in HeLa cells. The mean level of sperm Kglu was significantly reduced in A men compared with N men (P < 0.001) and was positively correlated with progressive motility (P < 0.001). The sodium glutarate-induced elevation of Kglu levels in A men with lower Kglu levels in sperm significantly improved the progressive motility (P < 0.001). Furthermore, the reduced sperm Kglu levels in A men was accompanied by an increase in sperm glutaryl-CoA dehydrogenase (a regulatory enzyme of Kglu). LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Although the present study indicated the involvement of sperm Kglu in maintaining progressive motility of human sperm, the underlying mechanism needs to be investigated further. WIDER IMPLICATIONS OF THE FINDINGS: The findings of this study provide an insight into the novel role of Kglu in human sperm and suggest that abnormality of sperm PLMs may be one of the causes of asthenozoospermia. STUDY FUNDING/COMPETING INTEREST(S): National Natural Science Foundation of China (81 771 644 to T.L.; 31 671 204 to X.Z. and 81 871 207 to H.C.); National Basic Research Program of China (973 Program, 2015CB943003 to X.Z.); Natural Science Foundation of Jiangxi, China (20171ACB21006 and 20161BAB204167 to T.L.; 20165BCB18001 to X.Z.). The authors have no conflicts of interest to declare.

Oncogenic role of microRNA-423-5p in hepatocellular carcinoma.

BACKGROUND: It has been found that microRNA-423-5p (miR423-5p) is an oncogenic factor and frequently upregulated in gastric carcinoma. However, the involvement of miR423-5p in hepatocellular carcinoma (HCC) has been rarely reported. The aim of this study was to assess whether miR423-5p is aberrantly expressed in HCC tissues, and to characterize its roles in the cancerous biology of HCC. METHODS: HCC and corresponding nonmalignant tissues were obtained from 115 patients during liver transplantation to detect the expression level of miR423-5p. The miR423-5p mimic and inhibitor were transfected into LM3 cell line. Cell viability assay, cell cycle analysis, transwell invasion and migration experiments were used to evaluate the oncogenic role of miR423-5p. RESULTS: miR423-5p was significantly upregulated in HCC compared with nonmalignant tissues, and this upregulation was negatively associated with recurrence-free survival. For patients beyond the Milan criteria, low expression of miR423-5p was correlated with better prognosis. Functional analysis showed that miR423-5p enhanced the proliferative, invasive and migratory capacity of HCC cells. CONCLUSIONS: miR423-5p contributed to the tumorigenesis and progression of HCC. It could be a new predictor in HCC patients beyond the Milan criteria and would help to improve patient outcomes and enlarge recipient pools of liver transplantation.

Overexpression of HOXA13 as a potential marker for diagnosis and poor prognosis of hepatocellular carcinoma.

HOXA13 is a member of homeobox genes that encode transcription factors regulating embryonic development and cell fate. Abnormal HOXA13 expression was reported in hepatocellular carcinoma (HCC), but its correlation with tumor angiogenesis and prognosis still remain unclear. This study was aimed to uncover the expression, diagnostic and prognostic significance of HOXA13 in HCC. Immunohistochemistry was performed to detect HOXA13 expression in HCC and corresponding paracarcinomatous tissues from 90 patients. Enzyme-linked immunosorbent assay was used to detect serum HOXA13 in 90 HCC patients and 20 healthy volunteers. Receiver operating characteristics was analyzed to calculate diagnostic accuracy of serum HOXA13, alpha-fetoprotein (AFP) and their combination. Immunoreactivity of HOXA13 was detected in 72.2% of HCC, and 12.2% of adjacent non-cancerous samples. HOXA13 expression was significantly associated with tumor size, microvascular invasion, pathological grade, tumor capsula status, AFP level, tumor-node-metastasis stage and positively correlated with VEGF (p < 0.001) and microvessel density (p < 0.001). The combination of serum HOXA13 and AFP had a markedly higher area under the curve than HOXA13 alone. HOXA13 expression was associated with unfavorable overall survival (OS) (p < 0.001) and disease-free survival (DFS) (p < 0.001). Multivariate analysis indicated that patients with HOXA13-expressing tumors had a significantly shorter OS (p = 0.030) and DFS (p = 0.005) than those with HOXA13-negative tumors. Thus, HOXA13 expression possibly plays an important role in tumor angiogenesis, progression and prognosis of HCC. Moreover, we demonstrate that serum HOXA13 may serve as a biomarker for early HCC diagnosing and predicting outcome.

Observation of complications assessed by Clavien-Dindo classification in different endoscopic procedures of benign prostatic hyperplasia: An observational study.

The Clavien-Dindo classification (CDC) was widely used in the assessment of surgical complications, but some inconsistencies always existed in urological literature. This study was aimed to report complications of the transurethral resection of the prostate (TURP), plasmakinetic resection of the prostate (PKRP), and holmium laser enucleation of the prostate (HoLEP) by using a more detailed way under the framework of CDC. A total of 623 eligible cases underwent endoscopic procedures from January 2018 and December 2020 were divided into the TURP group (212 cases), the PKRP group (208 cases), and the HoLEP group (203 cases) according to the surgical type. Patients' surgical complications assessed by the CDC were compared among the 3 groups. The operation time, intraoperative irrigation volume, postoperative irrigation time and volume, decrease in hemoglobin and sodium, postoperative catheterization time, visual analogue scale, hospital stay of the PKEP group and the HoLEP group were significantly less than those of the TURP group, and the decrease in hemoglobin and visual analogue scale in the HoLEP group were significantly lower than those in the PKEP group (all P < .05). The electrolyte disturbance, urinary tract irritation, and patients with grade II of CDC in the PKRP group were significantly lower than those in the TURP group; The electrolyte disturbance, lower abdominal pain, urinary tract irritation, intraoperative hemorrhage, secondary hemorrhage, clot retention, patients with grade I, II, III of CDC in the HoLEP group were significantly lower than those in the TURP group, and the urinary tract irritation, grade I, II of CDC in the HoLEP group was significantly lower than that in the PKRP group (all P < .05). The CDC should be recommended because of the enhanced insight into surgical complications, and the HoLEP should be given a priority for Benign prostatic hyperplasia (BPH) surgical treatment in terms of the merits in surgical characteristics and complications.

Glioblastoma cellular MAP4K1 facilitates tumor growth and disrupts T effector cell infiltration.

MAP4K1 has been identified as a cancer immunotherapy target. Whether and how cancer cell-intrinsic MAP4K1 contributes to glioblastoma multiforme (GBM) progression remains unclear. We found that MAP4K1 was highly expressed in the glioma cells of human GBM specimens. High levels of MAP4K1 mRNA were prevalent in IDH-WT and 1p/19q non-codeletion gliomas and correlated with poor prognosis of patients. MAP4K1 silencing inhibited GBM cell proliferation and glioma growth. Transcriptome analysis of GBM cells and patient samples showed that MAP4K1 modulated cytokine‒cytokine receptor interactions and chemokine signaling pathway, including IL-18R and IL-6R Importantly, MAP4K1 loss down-regulated membrane-bound IL-18R/IL-6R by inhibiting the PI3K-AKT pathway, whereas MAP4K1 restoration rescued this phenotype and therefore GBM cell proliferation. MAP4K1 deficiency abolished GBM cell pro-proliferation responses to IL-18, suggesting an oncogenic role of MAP4K1 via the intrinsic IL-18/IL-18R pathway. In addition, GBM cell-derived MAP4K1 impaired T-cell migration and reduced CD8+ T-cell infiltration in mouse glioma models. Together, our findings provide novel insight into the pathological significance of GBM cell-intrinsic MAP4K1 in driving tumor growth and immune evasion by remodeling cytokine-chemokine networks.

Angiotensin II type 2 receptor pharmacological agonist, C21, reduces the inflammation and pain hypersensitivity in mice with joint inflammatory pain.

Primary and secondary hyperalgesia develop in response to chronic joint inflammation due to peripheral and central mechanisms. Synovial macrophage and spinal microglia are involved in pain sensitization in arthritis. The level of angiotensin II type 2 receptor (AT2R) is related to the severity of arthritis. This study aimed to determine the role of AT2R in primary and secondary hyperalgesia in joint inflammatory pain in mice. After intra-articular CFA injection, primary hyperalgesia in the ipsilateral knee joint was measured by pressure application meter and gait analysis, secondary hypersensitivity in ipsilateral hind-paw was measured by von-Frey and Hargreaves tests following a combination of global AT2R-deficient (Agtr2-/-) mice and AT2R pharmacological agonist C21. Synovial macrophage and spinal microglia were collected for flow cytometry. Morphological reconstruction of microglia was detected by immunostaining. AT2R expression was investigated by quantitative polymerase chain reaction and western blot. Neuronal hyperactivity was evaluated by c-Fos and CGRP immunostaining. We found that pain hypersensitivity and synovial inflammation in Agtr2-/- mice were significantly exacerbated compared with wild-type mice; conversely, systemically administrated C21 attenuated both of the symptoms. Additionally, spinal microglia were activated, and an abundant increase of spinal AT2R was expressed on activated microglia in response to peripheral joint inflammation. Intrathecally-administrated C21 reversed the secondary hypersensitivity, accompanied by alleviation of spinal microglial activation, spinal neuronal hyperactivity, and calcitonin gene-related peptide content. These findings revealed a beneficial role of AT2R activating stimulation against pain hypersensitivity in joint inflammatory pain via direct modulation of synovial macrophage and spinal microglial activity.

Retrosplenial Cortex Effects Contextual Fear Formation Relying on Dysgranular Constituent in Rats.

Animal contextual fear conditioning (CFC) models are the most-studied forms used to explore the neural substances of posttraumatic stress disorder (PTSD). In addition to the well-recognized hippocampal-amygdalar system, the retrosplenial cortex (RSC) is getting more and more attention due to substantial involvement in CFC, but with a poor understanding of the specific roles of its two major constituents-dysgranular (RSCd) and granular (RSCg). The current study sought to identify their roles and underlying brain network mechanisms during the encoding processing of the rat CFC model. Rats with pharmacologically inactivated RSCd, RSCg, and corresponding controls underwent contextual fear conditioning. [18F]-fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG PET/CT) scanning was performed for each animal. The 5-h and 24-h retrieval were followed to test the formation of contextual memory. Graph theoretic tools were used to identify the brain metabolic network involved in encoding phase, and changes of nodal (brain region) properties linked, respectively, to disturbed RSCd and RSCg were analyzed. Impaired retrieval occurred in disturbed RSCd animals, not in RSCg ones. The RSC, hippocampus (Hip), amygdala (Amy), piriform cortex (Pir), and visual cortex (VC) are hub nodes of the brain-wide network for contextual fear memory encoding in rats. Nodal degree and efficiency of hippocampus and its connectivity with amygdala, Pir, and VC were decreased in rats with disturbed RSCd, while not in those with suppressed RSCg. The RSC plays its role in contextual fear memory encoding mainly relying on its RSCd part, whose condition would influence the activity of the hippocampal-amygdalar system.

Dexmedetomidine alleviates anxiety-like behavior in mice following peripheral nerve injury by reducing thehyperactivity of glutamatergic neurons in the anterior cingulate cortex.

BACKGROUND: Treatment of chronic pain is challenged by concurrent anxiety symptoms. Dexmedetomidine is known to produce sedation, analgesia, and anxiolysis. However, the neural mechanism of dexmedetomidine-elicited anxiolysis remains elusive. Here, we aimed to test the hypothesis that the anterior cingulate cortex might be involved in dexmedetomidine-induced anxiolysis in pain. METHODS: A common peroneal nerve ligation mouse model was used to test the dexmedetomidine-induced analgesia and anxiolysis by assessing mechanical allodynia, open-field, light-dark transition, and acoustic startle reflex tests. In vivo calcium signal fiber photometry and ex vivo whole-cell patch-clamp recordings were used to measure the excitability of glutamatergic neurons in anterior cingulate cortex. Modulation of glutamatergic neurons was performed by chemogenetic inhibition or activation via viral injection. RESULTS: Compared with vehicle, dexmedetomidine (4 µg/kg) alleviated mechanical allodynia (P < 0.001) and anxiety-like behaviors (P < 0.001). The glutamatergic neurons' excitability after dexmedetomidine administration was lower than that of the vehicle group (P = 0.001). Anxiety-like behaviors were rescued by inhibiting glutamatergic neurons in the model mice. Nociception-related anxiety-like behavior was induced by activation of glutamatergic neurons, which was rescued by dexmedetomidine. CONCLUSIONS: The reduction in glutamatergic neuronal activity in anterior cingulate cortex may be involved in dexmedetomidine-elicited anxiolysis in chronic pain.

Clemastine Ameliorates Myelin Deficits via Preventing Senescence of Oligodendrocytes Precursor Cells in Alzheimer's Disease Model Mouse.

Disrupted myelin and impaired myelin repair have been observed in the brains of patients and various mouse models of Alzheimer's disease (AD). Clemastine, an H1-antihistamine, shows the capability to induce oligodendrocyte precursor cell (OPC) differentiation and myelin formation under different neuropathological conditions featuring demyelination via the antagonism of M1 muscarinic receptor. In this study, we investigated if aged APPSwe/PS1dE9 mice, a model of AD, can benefit from chronic clemastine treatment. We found the treatment reduced brain amyloid-beta deposition and rescued the short-term memory deficit of the mice. The densities of OPCs, oligodendrocytes, and myelin were enhanced upon the treatment, whereas the levels of degraded MBP were reduced, a marker for degenerated myelin. In addition, we also suggest the role of clemastine in preventing OPCs from entering the state of cellular senescence, which was shown recently as an essential causal factor in AD pathogenesis. Thus, clemastine exhibits therapeutic potential in AD via preventing senescence of OPCs.

Shell-cross-linked micelles from PNIPAM-b-(PLL)2 Y-shaped miktoarm star copolymer as drug carriers.

Well-defined AB2 Y-shaped miktoarm star copolymers of PNIPAM-b-(PZLL)2 and PNIPAM-b-(PLL)2 were synthesized through the combination of atom transfer radical polymerization (ATRP), ring-opening polymerization (ROP), and click chemistry, where PNIPAM, PZLL, and PLL are poly(N-isopropylacrylamide), poly(epsilon-benzyloxy-carbonyl-L-lysine), and poly(L-lysine), respectively. Propargyl amine was employed as ROP initiator for the preparation of alkynyl-terminated PZLL. Diazide-terminated PNIPAM was obtained with an azide-containing ATRP initiator. The subsequent click reaction led to the formation of PNIPAM-b-(PZLL)2. After the removal of the benzyloxycarbonyl group, water-soluble PNIPAM-b-(PLL)2 was obtained. The core-shell micelles of PNIPAM-b-(PLL)2 were formed above lower critical solution temperature of PNIPAM block. At this temperature, the shell cross-linking was performed through the reaction between glutaraldehyde and the primary amine groups of the PLL shell, affording the micelles with the endurance to temperature and pH changes. These shell-cross-linked micelles were used as drug nanocarriers and the release profile was dually controlled by the solution temperature and the cross-linking degree.

Transplantation of GABAergic Interneuron Progenitor Attenuates Cognitive Deficits of Alzheimer's Disease Model Mice.

Excitatory (E) and inhibitory (I) balance of neural network activity is essential for normal brain function and of particular importance to memory. Disturbance of E/I balance contributes to various neurological disorders. The appearance of neural hyperexcitability in Alzheimer's disease (AD) is even suggested as one of predictors of accelerated cognitive decline. In this study, we found that GAD67+, Parvalbumin+, Calretinin+, and Neuropeptide Y+ interneurons were progressively lost in the brain of APP/PS1 mice. Transplanted embryonic medial ganglionic eminence derived interneuron progenitors (IPs) survived, migrated, and differentiated into GABAergic interneuron subtypes successfully at 2 months after transplantation. Transplantation of IPs hippocampally rescued impaired synaptic plasticity and cognitive deficits of APP/PS1 transgenic mice, concomitant with a suppression of neural hyperexcitability, whereas transplantation of IPs failed to attenuate amyloid-ß accumulation, neuroinflammation, and synaptic loss of APP/PS1 transgenic mice. These observations indicate that transplantation of IPs improves learning and memory of APP/PS1 transgenic mice via suppressing neural hyperexcitability. This study highlights a causal contribution of GABAergic dysfunction to AD pathogenesis and the potentiality of IP transplantation in AD therapy.

Fabrication and sensing application of a silver nanoshell array structure by a micro-flow injection method.

A two-dimensional periodic metallic spherical shell array structure with controllable geometric parameters was fabricated on the target substrate by microsphere templating and magnetron sputtering. The micro-flow injection method was used to prepare a two-dimensional colloidal microsphere template, and reactive ion etching (RIE) was used to change the spherical spacing. The geometric parameters and spectral characteristics of the spherical shell array structure were analyzed with the simulation software FDTD solutions. The nanostructural morphology and optical properties of the samples were characterized by scanning electron microscopy (SEM) and optical spectral analysis (OSA). The refractive index sensing application based on the principle of the local surface plasmon resonance (LSPR) and plasmonic interference was realized. The results show that the spherical shell arrays structure is sensitive to the surrounding environment, the refractive index sensitivity of spherical shell array structure is 527.07 nm per RIU and 922.25 nm per RIU, and the quality factor FOM is 16.5 and 15.3, respectively. The techniques demonstrated can produce large-area periodic nanostructure arrays with ultra-large production in cost-competitive ways. In addition, these properties make them applicable to multiple applications, such as surface plasmon sensors and various optical device.

Negative regulation of beta-adrenergic function by hydrogen sulphide in the rat hearts.

Beta-adrenoceptor is over-stimulated during myocardial ischemia, in which hydrogen sulphide (H2S) concentration was found to be lowered. The present study attempted to investigate if H2S modulates beta-adrenoceptor function and the underlying mechanism. We examined the effect of NaHS (a H2S donor) on myocyte contraction and electrically-induced (EI) intracellular calcium ([Ca2+](i)) transients upon beta-adrenergic stimulation in rat ventricular myocytes with a video edge tracker method and a spectrofluorometric method using fura-2/AM as a calcium indicator, respectively. We found that isoproterenol (ISO, 10(-9)-10(-6) M), a beta-adrenoceptor agonist, concentration-dependently increased the twitch amplitude of ventricular myocytes, which was attenuated by NaHS (10(-5)-10(-3) M) in a dose-dependent manner. The amplitudes and maximal velocities (+/-dl/dt) of myocyte twitch and EI-[Ca2+](i) transient amplitudes were enhanced by ISO, forskolin (an adenylyl cyclase activator), 8-bromoadenosine-3',5'-cyclic monophosphate (an activator of protein kinase A) and Bay K-8644 (a selective L-type Ca2+ channel agonist). Administration of NaHS (100 microM) only significantly attenuated the effects of ISO and forskolin. Moreover, NaHS reversed ISO-induced cAMP elevation and forskolin-stimulated adenylyl cyclase activity. In addition, stimulation of beta-adrenoceptor by ISO significantly decreased endogenous H2S production in rat ventricular myocytes. In conclusion, H2S may negatively modulate beta-adrenoceptor function via inhibiting adenylyl cyclase activity. Impairment of this negative modulation during ischemia may induce cardiac arrhythmias. Our study may provide a novel mechanism for ischemia-induced cardiac injury.

A single-center experience of sorafenib monotherapy in patients with advanced intrahepatic cholangiocarcinoma.

Patients with advanced intrahepatic cholangiocarcinoma (ICC) have a poor prognosis and the therapeutic options available for treating ICC are limited. Sorafenib, a multikinase inhibitor of vascular endothelial growth factor receptor 2 and 3, platelet derived growth factor receptor-ß, B-Raf proto-oncogene, serine/threonine kinase and C-Raf proto-oncogene, serine/threonine kinase, is a novel reference standard for the treatment of advanced hepatocellular carcinoma. Sorafenib has previously been demonstrated to exhibit significant antitumor activity in various cholangiocarcinoma cell lines and in xenograft ICC models. The present study aimed to assess the efficacy and safety of sorafenib as a single-agent treatment in patients with advanced ICC. Eligible patients were administere no prior therapy for metastatic or unresectable disease. Sorafenib was administered orally at a dose of 400 mg twice daily continuously. The primary endpoint was considered as the disease control rate (DCR) at 12 weeks. Secondary endpoints included time to progression (TTP), progression-free survival (PFS), overall survival (OS), duration of treatment (DOT) and the adverse event profile. A total of 15 patients were enrolled in the present study, with a median DOT of 3.2 months (range, 1.5-30 months). A total of 4 patients achieved a partial response and 7 patients achieved stable disease, with a DCR of 73.3%. The median OS time was 5.7 months [95% confidence interval (CI), 5.0-6.4], the PFS time was 5.5 months (95% CI, 3.9-7.1) and the median TTP was 3.2 months (range, 1.5-29 months). The most common toxicity was a skin rash, which w1as observed in 5 patients (33.3%). Grade 3 hand-foot syndrome was observed in 1 patient (6.7%), which required treatment termination. The results of the present study suggest that sorafenib monotherapy may exhibit promising anticancer activity in patients with advanced ICC and that it has a manageable toxicity profile.

Expression and prognostic significance of contactin 1 in human hepatocellular carcinoma.

BACKGROUND: CNTN1, a member of the CNTN family of neural cell-recognition molecules, is involved in tumor invasion and metastasis. Although the expression of CNTN1 has been reported in several human malignancies, the expression of CNTN1 in hepatocellular carcinoma (HCC) and its correlation with prognosis remain unclear. The aim of this study was to evaluate the expression of CNTN1 and determine the clinicopathological parameters and prognostic value of CNTN1 in HCC patients. MATERIALS AND METHODS: Quantitative real-time polymerase chain-reaction and Western blotting assays were performed to assess messenger RNA and protein levels of CNTN1 in 20 matched HCC specimens. The clinical and prognostic significance of CNTN1 in 90 cases of HCC was determined by immunohistochemistry. RESULTS: CNTN1 expression was higher in HCC compared to the expression found in adjacent tissues at both messenger RNA and protein levels (P<0.01). Notably, immunohistochemical results revealed that CNTN1 expression was significantly higher in HCC compared to adjacent tissues (54.4% vs 12.2%, P=0.01). Furthermore, positive CNTN1 expression was associated with tumor size, tumor capsulae, status of metastasis, and tumor-node-metastasis stage. Kaplan-Meier survival analysis showed that high CNTN1 was correlated with reduced overall survival (OS) rate (P<0.001) and disease-free survival (DFS) rate (P=0.001). Multivariate analysis identified CNTN1 as an independent poor prognostic factor of OS and DFS in HCC patients (P=0.007 and P=0.002, respectively). CONCLUSION: Our results suggest that CNTN1 could play an important role in HCC and serve as an independent unfavorable prognostic factor for OS and DFS and a potential therapeutic target for HCC.

Comb-Type Grafted Hydrogels of PNIPAM and PDMAEMA with Reversed Network-Graft Architectures from Controlled Radical Polymerizations.

Dual thermo- and pH-responsive comb-type grafted hydrogels of poly(N,N-dimethylaminoethyl methacrylate) (PDMAEMA) and poly(N-isopropylacrylamide) (PNIPAM) with reversed network-graft architectures were synthesized by the combination of atom transfer radical polymerization (ATRP), reversible addition-fragmentation chain transfer (RAFT) polymerization and click chemistry. Two kinds of macro-cross-linkers with two azido groups at one chain-end and different chain length [PNIPAM⁻(N3)2 and PDMAEMA⁻(N3)2] were prepared with N,N-di(ß-azidoethyl) 2-halocarboxylamide as the ATRP initiator. Through RAFT copolymerization of DMAEMA or NIPAM with propargyl acrylate (ProA) using dibenzyltrithiocarbonate as a chain transfer agent, two network precursors with different content of alkynyl side-groups [P(DMAEMA-co-ProA) and P(NIPAM-co-ProA)] were obtained. The subsequent azido-alkynyl click reaction of macro-cross-linkers and network precursors led to the formation of the network-graft hydrogels. These dual stimulus-sensitive hydrogels exhibited rapid response, high swelling ratio and reproducible swelling/de-swelling cycles under different temperatures and pH values. The influences of cross-linkage density and network-graft architecture on the properties of the hydrogels were investigated. The release of ceftriaxone sodium from these hydrogels showed both thermal- and pH-dependence, suggesting the feasibility of these hydrogels as thermo- and pH-dependent drug release devices.