A Robust Metal-Organic Framework with Scalable Synthesis and Optimal Adsorption and Desorption for Energy-Efficient Ethylene Purification.

Adsorptive separation is an energy-efficient alternative, but its advancement has been hindered by the challenge of industrially potential adsorbents development. Herein, a novel ultra-microporous metal-organic framework ZU-901 is designed that satisfies the basic criteria raised by ethylene/ethane (C2 H4 /C2 H6 ) pressure swing adsorption (PSA). ZU-901 exhibits an "S" shaped C2 H4 curve with high sorbent selection parameter (65) and could be mildly regenerated. Through green aqueous-phase synthesis, ZU-901 is easily scalable with 99 % yield, and it is stable in water, acid, basic solutions and cycling breakthrough experiments. Polymer-grade C2 H4 (99.51 %) could be obtained via a simulating two-bed PSA process, and the corresponding energy consumption is only 1/10 of that of simulating cryogenic distillation. Our work has demonstrated the great potential of pore engineering in designing porous materials with desired adsorption and desorption behavior to implement an efficient PSA process.

Outcome of Surgical Repair and Rehabilitation of Flexor Tendon Injuries in Zone II of the Hand: Systematic Review and Meta-Analysis.

PURPOSE: We performed a systematic review and meta-analysis to determine an optimal rehabilitation protocol following surgical repair for flexor tendon injury in zone II of the hand. METHODS: Records from PubMed, Embase, and Cochrane were retrieved from their establishment to January 12, 2020. Seven studies were included in the final analysis. A total of 569 digits with a flexor tendon injury in zone II of the hand were included in this meta-analysis: 135 in a place and hold group; 161 in an active flexion and extension group; and 273 in an early passive motion group. RESULTS: There was no significant difference between the place and hold and early passive motion regimes in the incidence of rupture. There was a significant difference between the active flexion and extension and early passive motion regimes in the incidence of rupture. In the early active motion group, the possibility of 1 or more grades of improvement on the Strickland grading system was increased. CONCLUSIONS: The early active motion group obtained greater total active motion than the early passive motion group. A higher risk of rupture was noted in the active flexion and extension subgroup repaired by 2-strand core suture. The 2-strand technique was not sufficient for active flexion and extension protocols. Further study in multistrand tendon repair technique with early active exercise in zone II should be undertaken to determine its efficacy. TYPE OF STUDY/LEVEL OF EVIDENCE: Therapeutic IV.

Hyperbaric Oxygen Improves the Survival and Angiogenesis of Fat Grafts after Autologous Fat Transplantation.

Objective: Currently, autologous fat transplantation (AFT) still has a low graft survival rate. Elevation of the AFT graft survival rate is a challenge. This study investigated the effect of hyperbaric oxygen (HBO) on AFT. Methods: Twelve adult male SD rats were randomly divided into two groups after AFT: the control group (n = 6) and the HBO group (n = 6). The rats were killed at 7, 14, and 28 days after transplantation to take the transplanted adipose tissues. The volume and weight of the tissues were detected. The pathological changes in the adipose tissues were observed after H&E staining. Microvessel density and levels of transforming growth factor- (TGF-) ß, tumor necrosis factor- (TNF-) α, and malondialdehyde (MDA) in the transplanted adipose tissues were measured with CD31 immunohistochemical stain, ELISA, and biochemical reagents, respectively. Additionally, the protein expression levels of vascular endothelial growth factor- (VEGF-) A and platelet-derived growth factor- (PDGF) A in the adipose tissues were detected by Western blot. Results: HBO significantly preserved the volume and weight of the transplanted adipose tissue (p < 0.01) and maintained the pathological structure of the transplanted adipose tissue. HBO therapy was effective in reducing inflammatory factor (TGF-ß and TNF-α) levels and oxidative stress (MDA) in the transplanted adipose tissue (p < 0.01) and significantly increased the level of CD31 and angiogenesis-related factors including VEGF-A and PDGF-A (p < 0.01) to promote angiogenesis. Conclusion: HBO therapy regulated the immune response of fat grafts, stimulated their angiogenesis, and ultimately promoted their survival after AFT.

Dacarbazine-Loaded Targeted Polymeric Nanoparticles for Enhancing Malignant Melanoma Therapy.

Dacarbazine (DTIC) dominates chemotherapy for malignant melanoma (MM). However, the hydrophobicity, photosensitivity, instability, and toxicity to normal cells of DTIC limit its efficacy in treating MM. In the present study, we constructed star-shaped block polymers nanoparticles (NPs) based on Cholic acid -poly (lactide-co-glycolide)-b-polyethylene glycol (CA-PLGA-b-PEG) for DTIC encapsulation and MM targeted therapy. DTIC-loaded CA-PLGA-b-PEG NPs (DTIC-NPs) were employed to increase the drug loading and achieve control release of DTIC, followed by further modification with nucleic acid aptamer AS1411 (DTIC-NPs-Apt), which played an important role for active targeted therapy of MM. In vitro, DTIC-NPs-Apt showed good pH-responsive release and the strongest cytotoxicity to A875 cells compared with DTIC-NPs and free DTIC. In vivo results demonstrated that the versatile DTIC-NPs-Apt can actively target the site of MM and exhibited excellent anti-tumor effects with no obvious side effects. Overall, this research provided multi-functional NPs, which endow a new option for the treatment of MM.

Current opinions on the mechanism, classification, imaging diagnosis and treatment of post-traumatic osteomyelitis.

Post-traumatic osteomyelitis (PTO) is a worldwide problem in the field of orthopaedic trauma. So far, there is no ideal treatment or consensus-based gold standard for its management. This paper reviews the representative literature focusing on PTO, mainly from the following four aspects: (1) the pathophysiological mechanism of PTO and the interaction mechanism between bacteria and the body, including fracture stress, different components of internal fixation devices, immune response, occurrence and development mechanisms of inflammation in PTO, as well as the occurrence and development mechanisms of PTO in skeletal system; (2) clinical classification, mainly the etiological classification, histological classification, anatomical classification and the newly proposed new classifications (a brief analysis of their scope and limitations); (3) imaging diagnosis, including non-invasive examination and invasive examination (this paper discusses their advantages and disadvantages respectively, and briefly compares the sensitivity and effectiveness of the current examinations); and (4) strategies, including antibiotic administration, surgical choices and other treatment programs. Based on the above-mentioned four aspects, we try to put forward some noteworthy sections, in order to make the existing opinions more specific.

Targeting Cell Death: Pyroptosis, Ferroptosis, Apoptosis and Necroptosis in Osteoarthritis.

New research has shown that the development of osteoarthritis (OA) is regulated by different mechanisms of cell death and types of cytokines. Therefore, elucidating the mechanism of action among various cytokines, cell death processes and OA is important towards better understanding the pathogenesis and progression of the disease. This paper reviews the pathogenesis of OA in relation to different types of cytokine-triggered cell death. We describe the cell morphological features and molecular mechanisms of pyroptosis, apoptosis, necroptosis, and ferroptosis, and summarize the current research findings defining the molecular mechanisms of action between different cell death types and OA.

Biomimetic and osteogenic 3D silk fibroin composite scaffolds with nano MgO and mineralized hydroxyapatite for bone regeneration.

Artificial bioactive materials have received increasing attention worldwide in clinical orthopedics to repair bone defects that are caused by trauma, infections or tumors, especially dedicated to the multifunctional composite effect of materials. In this study, a weakly alkaline, biomimetic and osteogenic, three-dimensional composite scaffold (3DS) with hydroxyapatite (HAp) and nano magnesium oxide (MgO) embedded in fiber (F) of silkworm cocoon and silk fibroin (SF) is evaluated comprehensively for its bone repair potential in vivo and in vitro experiments, particularly focusing on the combined effect between HAp and MgO. Magnesium ions (Mg2+) has long been proven to promote bone tissue regeneration, and HAp is provided with osteoconductive properties. Interestingly, the weak alkaline microenvironment from MgO may also be crucial to promote Sprague-Dawley (SD) rat bone mesenchymal stem cells (BMSCs) proliferation, osteogenic differentiation and alkaline phosphatase (ALP) activities. This SF/F/HAp/nano MgO (SFFHM) 3DS with superior biocompatibility and biodegradability has better mechanical properties, BMSCs proliferation ability, osteogenic activity and differentiation potential compared with the scaffolds adding HAp or MgO alone or neither. Similarly, corresponding meaningful results are also demonstrated in a model of distal lateral femoral defect in SD rat. Therefore, we provide a promising 3D composite scaffold for promoting bone regeneration applications in bone tissue engineering.

Effect of Irbesartan on Oxidative Stress and Serum Inflammatory Factors in Renal Tissues of Type 2 Diabetic Rats.

OBJECTIVE: To determine the effect of irbesartan on the activity of oxidative stress parameters of glutathione (GSH), superoxide enzyme (SOD) and malondialdehyde (MDA), and concentration of serum inflammatory factors of macrophage chemokine-1 (MCP-1), intercellular adhesion molecule-1 (ICAM-1) and tumor necrosis factor alpha (TNF-∝) in the renal tissue of type 2 diabetic rats. STUDY DESIGN: An experimental study. PLACE AND DURATION OF STUDY: The First People's Hospital of Ningyang County, Shandong Province, China, from April 2017 to March 2018. METHODOLOGY: Thirty healthy male Sprague Dawley rats were randomly divided into a normal control group, a diabetic model group, and an irbesartan treatment group, 10 in each group. Rat models of type 2 diabetes were prepared by highsugar high-fat diet with low-dose streptozotocin injection. Changes in blood glucose and lipids, serum GSH, SOD, MDA, MCP-1, ICAM-1, and TNF-∝ levels were determined. RESULTS: Levels of FBG, TC, TG and LDL, activity of GSH, SOD and MDA, and levels of serum MCP-1, ICAM-1 and TNF-∝ among three groups were statistically significant (all p < 0.001). Compared with the diabetic model group, FBG, TC, TG and LDL levels, MDA activity, MCP-1, ICAM-1 and TNF-∝ levels all decreased in rats of the irbesartan treatment group (all p <0.001), and both GSH and SOD activity increased (both p<0.001). CONCLUSION: Irbesartan can improve blood glucose and lipid levels in type 2 diabetic rats, and reduce renal damage by improving oxidative stress and inhibiting the release of inflammatory cytokines.

Ulk1 over-expression in human gastric cancer is correlated with patients' T classification and cancer relapse.

Ulk1 is a key autophagy protein. Here, we tested expression and potential function of Ulk1 in human gastric cancer. Ulk1 mRNA and protein were significantly elevated in multiple fresh human gastric cancer tissues. Its level was relatively low in surrounding normal epithelial tissues. Ulk1 over-expression was also observed in several gastric cancer cell lines (AGS, HGC-27, and SNU601). Remarkably, Ulk1 knockdown by targeted-shRNA inhibited AGS gastric cancer cell survival and proliferation. On the other hand, exogenous Ulk1 over-expression could further promote AGS cell survival and proliferation. Immunohistochemistry (IHC) staining assay of 145 paraffin-embedded gastric cancer tissues showed that Ulk1 was over-expressed in majority (114 out of 145) of gastric cancer tissues. Importantly, high Ulk1 expression in gastric cancer was correlated with patients' T classification and cancer relapse. Together, we demonstrate that Ulk1 over-expression in human gastric cancer is pro-survival. Its over-expression is associated with patients' T classification and cancer relapse.

Role of long non-coding RNA in tumor drug resistance.

Chemotherapy has been extensively used in tumor treatment, including either systemic or local treatment. Miserably, in many kinds of cancers, chemotherapy is gradually insensitive. The mechanisms of tumor drug resistance have been widely explored, yet have not been fully characterized. With several studies in the development of drug resistance, recent works have highlighted the involvement of non-coding RNAs in tumor development. A growing number of long non-coding RNAs (lncRNAs) have been identified as transcripts of larger than 200 nucleotides in length, which have low coding potential, but potentially coding small peptides with 50-70 amino acids. Despite so often being branded as transcriptional noise, it is becoming increasingly clear that a large number of lncRNAs are crucial molecular regulators of the processes of tumor involving the initiation and progression of human tumor. More recently, accumulating evidence is revealing an important role of lncRNA in tumor drug resistance and lncRNA expression profiling can be correlated with the evolution of tumor drug resistance. The long non-coding-RNA-mediated form of drug resistance brings yet another mechanism of drug resistance. So, exploiting the newly emerging knowledge of lncRNAs for the development of new therapeutic applications to overcome human tumor drug resistance will be significant.

Cerebrospinal fluid from rats given hypoxic preconditioning protects neurons from oxygen-glucose deprivation-induced injury.

Hypoxic preconditioning activates endogenous mechanisms that protect against cerebral ischemic and hypoxic injury. To better understand these protective mechanisms, adult rats were housed in a hypoxic environment (8% O2/92% N2) for 3 hours, and then in a normal oxygen environment for 12 hours. Their cerebrospinal fluid was obtained to culture cortical neurons from newborn rats for 1 day, and then the neurons were exposed to oxygen-glucose deprivation for 1.5 hours. The cerebrospinal fluid from rats subjected to hypoxic preconditioning reduced oxygen-glucose deprivation-induced injury, increased survival rate, upregulated Bcl-2 expression and downregulated Bax expression in the cultured cortical neurons, compared with control. These results indicate that cerebrospinal fluid from rats given hypoxic preconditioning protects against oxygen-glucose deprivation-induced injury by affecting apoptosis-related protein expression in neurons from newborn rats.

Correlation between advanced glycation end-products and the expression of fatty inflammatory factors in type II diabetic cardiomyopathy.

Diabetic cardiomyopathy (DCM) is one of the most severe complications of diabetes without a clear pathogenesis. Th is study investigated the adiponectin (APN) and leptin levels in type II DCM, as well as their correlation with advanced glycation end-products (AGEs). From 2011-2013, 78 type II diabetes mellitus (T2DM) cases (40-65 years old) in the Taian region were randomly selected. Based on the results of colour Doppler ultrasonography and coronary angiography, the cases were divided into a simple T2DM group (40 cases) and a DCM group (38 cases). Forty healthy subjects were used as normal control (NC). An enzyme-linked immunosorbent assay was performed to determine the levels of fa tty inflammatory factors such as APN, leptin and AGEs, and a correlation analysis was conducted. In the T2DM group, the APN levels were decreased but the leptin and AGE levels were significantly increased compared to the NC group. In the DCM group, the APN levels were decreased but the leptin and AGE levels were significantly increased (P<0.01) compared to the T2DM group. Th e AGE levels were positively correlated with disease progression and with fasting plasma glucose levels, glycated haemoglobin, insulin resistance and leptin, but were negatively correlated with APN levels. Additionally, the APN and leptin levels were independently related to the AGE levels. Fatty inflammatory factors play a significant role in the progression of both simple T2DM and DCM. Th e results of this study revealed the pathogenesis of DCM and indicated the potential significance of AGEs in DCM prevention and treatment.

Gabapentin Effects on PKC-ERK1/2 Signaling in the Spinal Cord of Rats with Formalin-Induced Visceral Inflammatory Pain.

Currently, the clinical management of visceral pain remains unsatisfactory for many patients suffering from this disease. While preliminary animal studies have suggested the effectiveness of gabapentin in successfully treating visceral pain, the mechanism underlying its analgesic effect remains unclear. Evidence from other studies has demonstrated the involvement of protein kinase C (PKC) and extracellular signal-regulated kinase1/2 (ERK1/2) in the pathogenesis of visceral inflammatory pain. In this study, we tested the hypothesis that gabapentin produces analgesia for visceral inflammatory pain through its inhibitory effect on the PKC-ERK1/2 signaling pathway. Intracolonic injections of formalin were performed in rats to produce colitis pain. Our results showed that visceral pain behaviors in these rats decreased after intraperitoneal injection of gabapentin. These behaviors were also reduced by intrathecal injections of the PKC inhibitor, H-7, and the ERK1/2 inhibitor, PD98059. Neuronal firing of wide dynamic range neurons in L6-S1 of the rat spinal cord dorsal horn were significantly increased after intracolonic injection of formalin. This increased firing rate was inhibited by intraperitoneal injection of gabapentin and both the individual and combined intrathecal application of H-7 and PD98059. Western blot analysis also revealed that PKC membrane translocation and ERK1/2 phosphorylation increased significantly following formalin injection, confirming the recruitment of PKC and ERK1/2 during visceral inflammatory pain. These effects were also significantly reduced by intraperitoneal injection of gabapentin. Therefore, we concluded that the analgesic effect of gabapentin on visceral inflammatory pain is mediated through suppression of PKC and ERK1/2 signaling pathways. Furthermore, we found that the PKC inhibitor, H-7, significantly diminished ERK1/2 phosphorylation levels, implicating the involvement of PKC and ERK1/2 in the same signaling pathway. Thus, our results suggest a novel mechanism of gabapentin-mediated analgesia for visceral inflammatory pain through a PKC-ERK1/2 signaling pathway that may be a future therapeutic target for the treatment of visceral inflammatory pain.

Repair of refractory wounds through grafting of artificial dermis and autologous epidermis aided by vacuum-assisted closure.

BACKGROUND: This study aimed to investigate the clinical efficacy of vacuum-assisted closure (VAC) combined with grafting of artificial dermis and autologous epidermis in the repair of refractory wounds. METHODS: Patients with refractory wounds underwent debridement. Then the VAC device was used to culture wound granulation tissue. After the wound granulation tissue began to grow, artificial dermis was grafted on the wounds with VAC treatment. Then autologous epidermis was grafted on the artificial dermis to repair the wounds after survival of the artificial epidermis. The study mainly observed length of the hospital stay, survival of the artificial dermis, time required for culture of the granulation tissue using VAC before grafting of the artificial dermis, survival time of the artificial dermis, survival conditions of the autologous epidermis, influence on functions of a healed wound at a functional part, healing conditions of donor sites, and recurrence conditions of the wounds. RESULTS: Healing was successful for 22 patients (95.7%), but treatment failed for 1 child. The 22 patients were followed up for 6 to 24 months. According to follow-up findings, the skin grafts had good color and a soft texture. They were wear resistant and posed no influence on function. The appearance of the final results was the same as that of the full-thickness skin graft. Mild or no pigmentation and no scar formation occurred at the donor sites, and the wounds did not recur. CONCLUSION: Vacuum-assisted closure combined with grafting of artificial dermis and autologous epidermis is an effective means for repairing refractory wounds and is worth clinical popularizing and application. LEVEL OF EVIDENCE V: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Antihyperglycemic and antihyperlipidemic activities of aqueous extract of Hericium erinaceus in experimental diabetic rats.

BACKGROUND: Hericium erinaceus, as a commonly used medicine or food, has attracted much attention due to its health effects when used as a home remedy for some diseases. The aim of this work was to investigate the hypoglycemic and hypolipidemic effects of aqueous extract of Hericium erinaceus (AEHE) in streptozotocin (STZ)-induced diabetic rats. METHODS: Diabetes was induced in Wistar rats by the administration of STZ (55 mg/kg BW.) intraperitoneally. AEHE (100 and 200 mg/kg BW.) was administered for a period of 28 days. The effects of AEHE on glucose, insulin, and lipid files in blood, and oxidative stress parameters in the liver were evaluated. The body weights of rats were recorded at day 0, 14 and 28th days. RESULTS: The administration of AEHE for 28 days in STZ diabetic rats resulted in a significant decrease in serum glucose level and a significant rise in serum insulin level. AEHE treatment attenuated lipid disorders. In addition, AEHE administration increased the activities of CAT, SOD, and GSH-Px, and GSH level, and reduced MDA level in the liver tissue significantly. CONCLUSION: Our results suggest that AEHE possesses hypoglycemic, hypolipidemic, and antioxidant properties in STZ-induced diabetes rats.

Elevated VEGF concentrations in ascites and serum predict adverse prognosis in ovarian cancer.

OBJECTIVE: To evaluate if serum and ascites VEGF-C concentration could be used as prognostic markers in ovarian cancer. MATERIALS AND METHODS: Serum and ascites VEGF-C were measured in 118 patients with ovarian cancer and 24 patients with benign gynaecological disease. ELISA method was employed to evaluate VEGF-C concentration. RESULTS: Serum and ascites in ovarian cancer patients were higher than those in benign gynaecological disease (P < 0.0001). Serum VEGF-C and ascites VEGF-C were both correlated with FIGO stage, tumor grade, and lymph node metastasis (P < 0.05). Univariate analysis revealed that FIGO stage (P < 0.0001), tumor grade (P < 0.0001), lymph node metastasis (P < 0.0001), serum VEGF-C concentration (P = 0.0001), and ascites VEGF-C concentration (P < 0.0001) were significantly correlated with overall survival. Including these variables in a multivariate analysis revealed that VEGF-C concentrations in serum and ascites were independent predictors of shorter overall survival. CONCLUSION: Elevated VEGF-C concentrations in serum and ascites can be seen as the novel, widely available independent predictors of shorter overall survival in ovarian cancer.

[Pathomorphological observation of the hypertrophic scar induced by injury to conical structure in female red Duroc pig].

UNLABELLED: To further explore the relationship between hypertrophic scar and injury to conical structure of skin and the pathogenesis of hypertrophic scar, and to reproduce an optimal animal model of hypertrophic scar. METHODS: The back of two FRDP pigs were shaved, and a piece of normal skin was harvested for the observation of conical structure of skin. Skin wounds with depth of 0.38 mm, 0.76 mm, 1.14 mm and 1.52 mm, respectively, were created by gas-driven dermatome. Eight wounds measuring 7.0 cm x 7.0 cm were created on each pig. The wounds were divided into 4 groups according to the wound depth with 4 wounds in each group, i.e. 0.38 mm group, 0.76 mm group, 1.14 mm group and 1.52 mm group. The 0.38 mm and 0.76 mm groups were designated as superficial wound groups and 1.14 mm and 1.52 mm groups as deep wound groups. The wounds were allowed to heal without treatment. Tissue samples from the wound were harvested on 0, 10, 30, 60, 90 and 150 post injury day (PID) , and they were sectioned for HE staining and staining for elastic fibers (VVG). The wound healing and the scar formation were observed with naked eye. The skin conical structures in normal and injured skin were also observed. The morphology of hypertrophic scar was observed, and the thickness of the scar tissue was determined and scored. RESULTS: The wounds in superficial wound groups healed within 3 weeks with flat surface without scar formation. The wounds in deep wound groups healed later than 4 weeks with thick, hairless, hard in texture, with depigmentation or pigmentation, finally forming contracture. The skin conical structure could be found on the back of FRDP with HE and VVG staining, and it was similar to that of human in terms of the structure. In superficial wound groups, the upper part of the skin conical structure was injured, but fat fornix and glands were intact. In deep wound groups, the lower part of the skin cone, together with the fat fornix and gland were all injured. On the 150th post injury day, the histological picture of the tissue in superficial wound groups was similar to that of normal skin. But the skin conical structure could not be found in deep wound groups, and the wounds were filled by a large accumulation of disarrayed and irregularly arranged collagen fibers. With passage of time, the scar became thicker and thicker, and the scar hypertrophy reached the zenith in 150th PID. CONCLUSION: The injury of skin conical structure can lead to the formation of hypertrophic scar. FRDP can be used to reproduce and ideal model of hypertrophic scar.

Effects of various muscarinic ligands on M2AChR-Gi1alpha fusion protein expressed in Sf9 insect cells.

AIM: To generate M2AChR-Gi1alpha fusion protein in baculovirus-Sf9 cells system and detect the effects of various muscarinic ligands and magnesium ion on the interaction of fused M2AChR and Gi1alpha. METHODS: M2AChR-Gi1alpha fused DNA was generated in a two step polymerase chain reaction (PCR) and then expressed in Sf9 cells to produce fusion protein. [3H] L-quinuclidinyl benzilate (QNB) and 35S GTPgammaS binding experiments were performed to study the function of M2AChR-Gi1alpha fusion protein. RESULTS: The expression level of M2AChR-Gi1alpha was (20.12 +/- 0.14) nmol/g protein. The affinity of GDP to Gi1alpha partner changed in the presence of different muscarinic ligands. IC50 values (95 % confidence limit) of GDP in the presence of acetylcholine (ACh), carbamylcholine, (4-hydroxy-2-butynyl)-1-trimethylammonium-m-chloro-carbanilate chloride (McN-A-343), pilocarpine, and atropine were 178 (148 - 214) micromol/L, 158 (126 - 199) micromol/L, 66 (56 - 78) micromol/L, 62 (55 -7 2) micromol/L, and 5.0 (4.6 - 5.5) micromol/L, respectively, and that in the absence of muscarinic ligand was 15.9 (14.3 - 17.6) micromol/L. Apparent affinity for GDP in the presence of carbamylcholine was markedly decreased with increasing MgCl2 concentrations, although the apparent affinity in the presence of atropine was not affected. CONCLUSION: The M2AChR-Gi1alpha fusion protein has the pharmacological specificity of M2 receptor and the efficient signaling of the two partners. Affinity of GDP to ligand-bound fusion protein represents the species of muscarinic ligands. Mg2+ is necessary for the action of M2AChR on Gi1alpha.