Characterization of Point Mutations in the cdtA Gene of the Cytolethal Distending Toxin of Actinobacillus Actinomycetemcomitans

The Cdt is a family of gram-negative bacterial toxins that typically arrest eukaryotic cells in the G0/G1 or G2/M phase of the cell cycle. The toxin is a heterotrimer composed of the cdtA, cdtB and cdtC gene products. Although it has been shown that the CdtA protein subunit binds to cells in culture and in an enzyme-linked immunosorbent assay (CELISA) the precise mechanisms by which CdtA interacts with CdtB and CdtC has not yet been clarified. In this study we employed a random mutagenesis strategy to construct a library of point mutations in cdtA to assess the contribution of individual amino acids to binding activity and to the ability of the subunit to form biologically active holotoxin. Single unique amino acid substitutions in seven CdtA mutants resulted in reduced binding of the purified recombinant protein to Chinese hamster ovary cells and loss of binding to the fucose-containing glycoprotein, thyroglobulin. These mutations clustered at the 5′- and 3′-ends of the cdtA gene resulting in amino acid substitutions that resided outside of the aromatic patch region and a conserved region in CdtA homologues. Three of the amino acid substitutions, at positions S165N (mutA81), T41A (mutA121) and C178W (mutA221) resulted in gene products that formed holotoxin complexes that exhibited a 60% reduction (mutA81) or loss (mutA121, mutA221) of proliferation inhibition. A similar pattern was observed when these mutant holotoxins were tested for their ability to induce cell cycle arrest and to convert supercoiled DNA to relaxed and linear forms in vitro. The mutations in mutA81 and mutA221 disrupted holotoxin formation. The positions of the amino acid substitutions were mapped in the Haemophilus ducreyi Cdt crystal structure providing some insight into structure and function

Effect of the of remazolam toluenesulfonate and IPACK on postoperative rehabilitation and sedation in elderly patients undergoing knee arthroplasty

Purpose: To investigate the effect of the combination of remazolam toluenesulfonate and infiltration between the popliteal artery and capsule of the knee (IPACK) on the rehabilitation and sedation of elderly patients undergoing knee arthroplasty. Methods: A total of 84 elderly patients who underwent knee arthroplasty in Quzhou Affiliated Hospital of Wenzhou Medical University between January 2021 and December 2021 were given IPACK in combination with adductor canal block under ultrasound guidance. Forty (40) patients were given anesthesia with propofol + sufentanil + cis-benzene sulfonic acid atracurium (control group), while 44 patients received anesthesia with remazolam toluenesulfonate + sufentanil + cis-benzene sulfonic acid atracurium (study group). Various clinical indices in the patients were assessed Results: Relative to control, there were significantly lower visual analog scale (VAS) scores at 6 and 12 h after operation, lower frequency of post-operation pumping with PCA pump, lower frequency of remedial analgesia within 48 h after operation (p < 0.05). Moreover, the time taken for spontaneous recovery of breathing, recovery time, and extubation time were similar in the two groups (p > 0.05). The muscle strength of quadriceps femoris of the affected knee joint was higher in the study group than in the control group at 12, 24, and 48 h, after operation (p < 0.05). Conclusion: The combination of remazolam toluenesulfonate with IPACK effectively alleviate pain in elderly patients undergoing knee arthroplasty and improves sedation without increasing cognitive dysfunction. This treatment strategy has potentials for further investigation for possible large-scale application

Characterization of Point Mutations in the cdtA Gene of the Cytolethal Distending Toxin of Actinobacillus Actinomycetemcomitans

The Cdt is a family of gram-negative bacterial toxins that typically arrest eukaryotic cells in the G0/G1 or G2/M phase of the cell cycle. The toxin is a heterotrimer composed of the cdtA, cdtB and cdtC gene products. Although it has been shown that the CdtA protein subunit binds to cells in culture and in an enzyme-linked immunosorbent assay (CELISA) the precise mechanisms by which CdtA interacts with CdtB and CdtC has not yet been clarified. In this study we employed a random mutagenesis strategy to construct a library of point mutations in cdtA to assess the contribution of individual amino acids to binding activity and to the ability of the subunit to form biologically active holotoxin. Single unique amino acid substitutions in seven CdtA mutants resulted in reduced binding of the purified recombinant protein to Chinese hamster ovary cells and loss of binding to the fucose-containing glycoprotein, thyroglobulin. These mutations clustered at the 5′- and 3′-ends of the cdtA gene resulting in amino acid substitutions that resided outside of the aromatic patch region and a conserved region in CdtA homologues. Three of the amino acid substitutions, at positions S165N (mutA81), T41A (mutA121) and C178W (mutA221) resulted in gene products that formed holotoxin complexes that exhibited a 60% reduction (mutA81) or loss (mutA121, mutA221) of proliferation inhibition. A similar pattern was observed when these mutant holotoxins were tested for their ability to induce cell cycle arrest and to convert supercoiled DNA to relaxed and linear forms in vitro. The mutations in mutA81 and mutA221 disrupted holotoxin formation. The positions of the amino acid substitutions were mapped in the Haemophilus ducreyi Cdt crystal structure providing some insight into structure and function

Characterization of Point Mutations in the \u3cem\u3ecdtA\u3c/em\u3e Gene of the Cytolethal Distending Toxin of \u3cem\u3eActinobacillus Actinomycetemcomitans\u3c/em\u3e

The Cdt is a family of gram-negative bacterial toxins that typically arrest eukaryotic cells in the G0/G1 or G2/M phase of the cell cycle. The toxin is a heterotrimer composed of the cdtA, cdtB and cdtC gene products. Although it has been shown that the CdtA protein subunit binds to cells in culture and in an enzyme-linked immunosorbent assay (CELISA) the precise mechanisms by which CdtA interacts with CdtB and CdtC has not yet been clarified. In this study we employed a random mutagenesis strategy to construct a library of point mutations in cdtA to assess the contribution of individual amino acids to binding activity and to the ability of the subunit to form biologically active holotoxin. Single unique amino acid substitutions in seven CdtA mutants resulted in reduced binding of the purified recombinant protein to Chinese hamster ovary cells and loss of binding to the fucose-containing glycoprotein, thyroglobulin. These mutations clustered at the 5′- and 3′-ends of the cdtA gene resulting in amino acid substitutions that resided outside of the aromatic patch region and a conserved region in CdtA homologues. Three of the amino acid substitutions, at positions S165N (mutA81), T41A (mutA121) and C178W (mutA221) resulted in gene products that formed holotoxin complexes that exhibited a 60% reduction (mutA81) or loss (mutA121, mutA221) of proliferation inhibition. A similar pattern was observed when these mutant holotoxins were tested for their ability to induce cell cycle arrest and to convert supercoiled DNA to relaxed and linear forms in vitro. The mutations in mutA81 and mutA221 disrupted holotoxin formation. The positions of the amino acid substitutions were mapped in the Haemophilus ducreyi Cdt crystal structure providing some insight into structure and function

Effects of ultrasound-guided brachial plexus block in combination with sevoflurane general anesthesia via laryngeal mask on upper limb tumor and fracture surgery for children based on inflammatory stress response

Sevoflurane is used successfully for general anesthesia via laryngeal mask on upper limb tumors like osteosarcoma and Ewings sarcoma and fractures in orthopedic care centers. With the advancement in interventional radiology, many procedures are now available to support surgeons to carry out such procedures without complications. In this study, we tried to evaluate the effects of ultrasound-guided brachial plexus block in combination with sevoflurane general anesthesia via laryngeal mask for upper limb tumor (osteosarcoma in the metaphysic of long bones of upper limb and Ewing sarcoma-second most malignant tumor in children) and fracture surgery in children based on inflammatory stress response. Eighty-six children who were admitted to undergo surgery for upper limb tumor and fractures from June 2018 to November 2019 were selected and divided into control and study groups according to anesthesia methods (n= 43). The control group received only sevoflurane superficial general anesthesia via laryngeal mask, besides which the study group received ultrasound-guided brachial plexus block. The surgical, anesthetic outcomes and adverse reactions were compared. Pain was assessed by the visual analogue scale (VAS) 4 h, 8 h and 12 h after surgery. The serum levels of interleukin-1β (IL-1β), IL-10, C-reactive protein (CRP), tumor necrosis factor-α (TNF-α), epinephrine (E), norepinephrine (NE), cortisol (Cor) and insulin (Ins) were measured before surgery, on the day of surgery and 3 days after surgery, respectively. Compared with the control group, the surgical time and recovery time of the study group were shorter, the pain occurred later, the intraoperative respiration was slower, and the incidence rate of adverse reactions was significantly lower. The VAS scores of the study group were significantly lower than those of the control group 4 h, 8 h and 12 h after surgery. The IL-1β, IL-10, CRP, TNF-α, E, NE, Cor, and Ins levels of the study group were also significantly lower than those of the control group on the day of surgery and 3 days after surgery. Observations of this study suggest that sevoflurane superficial general anesthesia via laryngeal mask in combination with brachial plexus block can be used in the upper limb tumor removal and fracture surgery for children. This strategy has the advantages of short recovery time, few adverse reactions, obvious sedative and analgesic effects, and low inflammatory stress response

Effects of ultrasound-guided brachial plexus block in combination with sevoflurane general anesthesia via laryngeal mask on upper limb tumor and fracture surgery for children based on inflammatory stress response

734-739Sevoflurane is used successfully for general anesthesia via laryngeal mask on upper limb tumors like osteosarcoma and Ewings sarcoma and fractures in orthopedic care centers. With the advancement in interventional radiology, many procedures are now available to support surgeons to carry out such procedures without complications. In this study, we tried to evaluate the effects of ultrasound-guided brachial plexus block in combination with sevoflurane general anesthesia via laryngeal mask for upper limb tumor (osteosarcoma in the metaphysic of long bones of upper limb and Ewing sarcoma-second most malignant tumor in children) and fracture surgery in children based on inflammatory stress response. Eighty-six children who were admitted to undergo surgery for upper limb tumor and fractures from June 2018 to November 2019 were selected and divided into control and study groups according to anesthesia methods (n= 43). The control group received only sevoflurane superficial general anesthesia via laryngeal mask, besides which the study group received ultrasound-guided brachial plexus block. The surgical, anesthetic outcomes and adverse reactions were compared. Pain was assessed by the visual analogue scale (VAS) 4 h, 8 h and 12 h after surgery. The serum levels of interleukin-1β (IL-1β), IL-10, C-reactive protein (CRP), tumor necrosis factor-α (TNF-α), epinephrine (E), norepinephrine (NE), cortisol (Cor) and insulin (Ins) were measured before surgery, on the day of surgery and 3 days after surgery, respectively. Compared with the control group, the surgical time and recovery time of the study group were shorter, the pain occurred later, the intraoperative respiration was slower, and the incidence rate of adverse reactions was significantly lower. The VAS scores of the study group were significantly lower than those of the control group 4 h, 8 h and 12 h after surgery. The IL-1β, IL-10, CRP, TNF-α, E, NE, Cor, and Ins levels of the study group were also significantly lower than those of the control group on the day of surgery and 3 days after surgery. Observations of this study suggest that sevoflurane superficial general anesthesia via laryngeal mask in combination with brachial plexus block can be used in the upper limb tumor removal and fracture surgery for children. This strategy has the advantages of short recovery time, few adverse reactions, obvious sedative and analgesic effects, and low inflammatory stress response

Spatial variation in grain-size population of surface sediments from northern Bering Sea and western Arctic Ocean: implications for provenance and depositional mechanisms

In general, sediments in nature comprise populations of various diameters. Accurate information regarding the sources and depositional mechanisms of the populations can be obtained through their temporal and spatial comparisons. In this study, the grain size distribution of surface sediments from the Bering Sea and western Arctic Ocean were fitted and partitioned into populations using a log-normal distribution function. The spatial variations in the populations indicate differences in their sources and deposition mechanisms. The sediments on most of the Bering Sea Shelf originated from the Yukon River, and were transported westward by waves and currents. However, the presence of a coarser population outside Anadyr Bay was the result of Anadyr River transport. Additionally, a northward transport trend of fine suspended particles was observed on the west side of the Bering Sea Shelf. The sediments in Hope Valley in the south Chukchi Sea also originated from the Yukon River. The coarser population on the central Chukchi Sea Shelf originated from coast of Alaska to the east, not the Yukon River, and was transported by sea ice and bottom brine water. The populations of sediments from the Chukchi Basin and the base of the Chukchi Sea Slope are the result of sea ice and eddy action. Surface sediments from the western high Arctic Ocean predominantly comprised five populations, and two unique populations with mode diameters of 50–90 μm and 200–400 μm, respectively, were ubiquitous in the glacial and interglacial sediments. It was difficult to distinguish whether these two populations originated from sea ice or icebergs. Therefore, caution should be exercised when using either the > 63 μm or > 250 μm fractions in sediments as a proxy index for iceberg and ice sheet variation in the high Arctic Ocean