Comparison of antibiotic-impregnated bone cement coverage versus vacuum sealing drainage in semi-open bone grafting using for tibial fracture with infected bone and soft tissue defect: a retrospective analysis.

OBJECTIVE: To compare antibiotic-impregnated bone cement coverage (bone cement surface technique; BCS-T) versus vacuum sealing drainage (VSD) for tibial fracture with infected bone and soft tissue defect. METHOD: This retrospective analysis compared the clinical outcomes in patients undergoing BCS-T (n = 16) versus VSD (n = 15) for tibial fracture with infected bone and soft tissue defect at the Third Hospital of Hebei Medical University from March 2014 to August 2019. For BCS-T group, osseous cavity was filled with autograft bone graft after debridement, and then the wound was covered with a 3-mm layer of bone cement impregnated with vancomycin and gentamycin. The dressing was changed every day in the first week, and every 2 ~ 3 days in the second week. For VSD group, a negative pressure of -150 ~ -350 mmHg was maintained, and the dressing was changed every 5-7 days. All patients received antibiotics treatment based on bacterial culture results for 2 weeks. RESULTS: The 2 groups did not differ in age, sex and key baseline characteristics, including type of Gustilo-Anderson classification, size of the bone and soft tissue defect, the percentage of primary debridement, bone transport, and the time from injury to bone grafting. The median follow-up was 18.9 months (range:12-40). The time to complete coverage of bone graft by granulation tissue was 21.2 (15.0-44.0) and 20.3 (15.0-24.0) days in the BCS-T and VSD groups, respectively (p = 0.412). The 2 groups also did not differ in wound healing time (3.3 (1.5-5.5) versus 3.2(1.5-6.5) months; p = 0.229) and bone defect healing time (5.4(3.0-9.6) versus 5.9(3.2-11.5) months; p = 0.402). However, the cost of covering material was significantly reduced in the BCS-T group (2071 ± 134 versus 5542 ± 905 yuan; p = 0.026). Paley functional classification at 12 months did not differ between the 2 groups (excellent in 87.5% versus 93.3% in the 2 groups; p = 0.306). CONCLUSION: BCS-T could achieve clinical outcomes similar to VSD in patients receiving bone graft for tibial fracture with infected bone and soft tissue defect, but material cost was significantly reduced. Randomized controlled trials are needed to verify our finding.

A sensitive, expandable AQC-based LC-MS/MS method to measure amino metabolites and sphingolipids in cell and serum samples.

Sphingolipids are a major lipid species found in all eukaryotes. Among structurally complex and diversified lipids, sphingoid bases have been heavily linked to various metabolic diseases. However, most current LC-MS-based methods lack the sensitivity to detect low-abundant sphingoid bases. The 6-Aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) derivatization reagent, which efficiently forms covalent bonds with amino groups, has been widely used for amino acid detection. Nevertheless, the commonly used reverse-phase HPLC method for amino acid analysis is not suitable for amphipathic sphingolipids. To address this issue, we report a robust reverse-phase HPLC-MS/MS method capable of separating and detecting hydrophilic amino acids and sphingoid bases in a single run with high sensitivity. This method is also inclusive of other amino metabolites with an expandable target list. We tested this method under various conditions and samples, demonstrating its high reproducibility and sensitivity. Using this approach, we systematically analyzed human serum samples from healthy individuals, dyslipidemia, and type II diabetes mellitus (T2DM) patients, respectively. Two sphingolipids and five amino acids were identified with significant differences between the control and T2DM groups, highlighting the potential of this method in clinical studies.

CYP1B1 affects the integrity of the blood-brain barrier and oxidative stress in the striatum: An investigation of manganese-induced neurotoxicity.

AIMS: Excessive influx of manganese (Mn) into the brain across the blood-brain barrier induces neurodegeneration. CYP1B1 is involved in the metabolism of arachidonic acid (AA) that affects vascular homeostasis. We aimed to investigate the effect of brain CYP1B1 on Mn-induced neurotoxicity. METHOD: Brain Mn concentrations and α-synuclein accumulation were measured in wild-type and CYP1B1 knockout mice treated with MnCl2 (30 mg/kg) and biotin (0.2 g/kg) for 21 continuous days. Tight junctions and oxidative stress were analyzed in hCMEC/D3 and SH-SY5Y cells after the treatment with MnCl2 (200 µM) and CYP1B1-derived AA metabolites (HETEs and EETs). RESULTS: Mn exposure inhibited brain CYP1B1, and CYP1B1 deficiency increased brain Mn concentrations and accelerated α-synuclein deposition in the striatum. CYP1B1 deficiency disrupted the integrity of the blood-brain barrier (BBB) and increased the ratio of 3, 4-dihydroxyphenylacetic acid (DOPAC) to dopamine in the striatum. HETEs attenuated Mn-induced inhibition of tight junctions by activating PPARγ in endothelial cells. Additionally, EETs attenuated Mn-induced up-regulation of the KLF/MAO-B axis and down-regulation of NRF2 in neuronal cells. Biotin up-regulated brain CYP1B1 and reduced Mn-induced neurotoxicity in mice. CONCLUSIONS: Brain CYP1B1 plays a critical role in both cerebrovascular and dopamine homeostasis, which might serve as a novel therapeutic target for the prevention of Mn-induced neurotoxicity.

Human Retinal Pigment Epithelial Cells Overexpressing the Neuroprotective Proteins PEDF and GM-CSF to Treat Degeneration of the Neural Retina.

BACKGROUND: Non-viral transposon-mediated gene delivery can overcome viral vectors' limitations. Transposon gene delivery offers the safe and life-long expression of genes such as Pigment Epithelium-Derived Factor (PEDF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) to counteract retinal degeneration by reducing oxidative stress damage. OBJECTIVE: The study aimed at using Sleeping Beauty transposon to transfect human Retinal Pigment Epithelial (RPE) cells with the neuroprotective factors PEDF and GM-CSF to investigate the effect of these factors on oxidative stress damage. METHODS: Human RPE cells were transfected with PEDF and GM-CSF by electroporation, using the hyperactive Sleeping Beauty transposon gene delivery system (SB100X). Gene expression was determined by RT-qPCR, and protein level by Western Blot as well as ELISA. The cellular stress level and the neuroprotective effect of the proteins were determined by measuring the concentrations of the antioxidant glutathione in human RPE cells, and conducting immunohistochemical examination of retinal integrity, inflammation, and apoptosis of rat Retina-Organotypic Cultures (ROC) exposed to H2O2. RESULTS: Human RPE cells were efficiently transfected showing a significantly augmented gene expression and protein secretion. Human RPE cells overexpressing PEDF and/or GM-CSF or pretreated with recombinant proteins presented significantly increased glutathione levels post- H2O2 incubation than non-transfected/untreated controls. rPEDF and/or rGM-CSF-treated ROC exhibited decreased inflammatory reactions and cell degeneration. CONCLUSION: GM-CSF and/or PEDF could be delivered successfully to RPE cells with combined use of SB100X and electroporation. PEDF and/or GM-CSF reduced H2O2-mediated oxidative stress damage in RPE cells and ROC offering an encouraging technique to re-establish a cell protective environment to halt age-related retinal degeneration.

Characterization of peptidoglycan hydrolase in Cag pathogenicity island of Helicobacter pylori.

The Cag Type IV secretion apparatus proteins in Helicobacter pylori can mediate the injection of effector CagA protein into eukaryotic target cells. Although this apparatus forms an important pathway for bacterium-host interaction, its assembly process in vivo is poorly understood, and the proteins which contribute to break the bacterial cell walls in Cag-PAI have not yet been identified. The cagγ gene in Cag-PAI is a unique member that contains a conserved SLT catalysis domain, which makes it an attracting question whether cagy gene has the capacity to digest the bacterial cell wall. In the current study, therefore, the cagγ gene was cloned from the H. pylori NCTC 11637 and expressed in Escherichia coli, and its lytic effect on cell walls in vitro was observed. Results indicated that Cagγ protein has a lytic activity against bacterial cell walls. An allelic-exchange mutant (Δcagγ) was further constructed to investigate the relationship between Cagγ and effector CagA translocation. These results suggested that Cagγ contributed to the assembly of Cag Type IV secretion apparatus by digesting the peptidoglycan meshwork of bacterial cell walls.

Intertrochanteric valgus osteotomy for post-traumatic coxa vara after proximal femur fractures: A retrospective study.

ABSTRACT: To investigate the clinical effects of a new intertrochanteric valgus osteotomy technique designed by the authors for treatment of post-traumatic coxa varus after proximal femur fractures. Retrospectively analyzed 11 patients who developed coxa vara after sustaining proximal femoral fractures were treated with intertrochanteric valgus osteotomy from December 2005 to December 2018 in our hospital. This study included 6 cases of intertrochanteric fracture deformity union, 3 cases of subtrochanteric fracture nonunion and 2 cases of femoral neck fracture nonunion. Measured the degree of coxa varus, the differences in the lower limb length and force line in all patients. Evaluated hip function with the Harris hip score. All injuries were treated with the authors' intertrochanteric valgus osteotomy technique. The average follow-up period was 3 years and evaluated the clinical effects by radiological examination and the Harris hip score. The average neck-shaft angle increased 35.0° (99.1°-134.1°) and the average limb shortening lengthened 1.9 cm (2.9-1.0 cm) after surgery. The average operating time was 67.2 minutes and blood loss was 237.7 ml. The osteotomy position healed in all patients 3 months later. Union of the 2 old femoral neck fractures was achieved 4 and 6 months after surgery, respectively, and no femoral head necrosis occurred during follow-up. The Harris hip score increased an average of 49 points (44.1-93.1 points) at 1 year postoperatively. Our self-designed intertrochanteric valgus osteotomy technique showed a favorable clinical effect to treatment coxa vara and can be used in the clinical setting.

[Autofluorescence combined with spectral domain optical coherence tomography for diagnosis and follow-up of acute Vogt-Koyanagi-Harada disease].

OBJECTIVE: To evaluate the value of fundus autofluorescence (FAF) imaging combined with spectral domain optical coherence tomography (SD-OCT) in diagnosis, prognostic assessment and follow-up observation of acute Vogt-KoyanagiHarada (VKH) disease. METHODS: Clinical data were collected from 12 patients (23 eyes) with acute VKH disease treated in our hospital from May, 2018 to November, 2019, including detailed medical history, best corrected visual acuity (BCVA), and results of slit lamp biomicroscopy, fundus photography, SD-OCT, fundus fluorescein angiography (FFA) and FAF imaging.SDOCT and FAF imaging were repeated after a course of treatment and in follow-up examination, and the results were compared with those at the time of admission. RESULTS: VKH disease involved both eyes in 11 patients (91.7%).Fundus photography showed optic disc edema in 16 eyes (69.6%), and multiple retinal neuroepithelial detachment was detected by SD-OCT in all the involved eyes (100%).IN all the eyes, FFA revealed small and dense fluorescein leakage in the early stage and fluorescein accumulation in advanced stages of VHK disease to form multiple dye pooling in the areas of serous detachment.Hyperauto fluorescence was a common finding in FAF imaging (100%), and the area involved was consistent with that of fluorescein accumulation shown by FAF imaging.Ten eyes (43.5%) showed patches of relative hypoautofluorescence in the hyperauto fl uorescence areas, and granular hyperauto fl uorescence was found in the lesions in 4 eyes (17.4%).During the remission period of VKH disease, FAF imaging showed normal finding in 8 eyes (34.8%) and reduced areas (by 55.2%) and intensity (by 46.5%) of hyperautofluorescence in 9 eyes (39.1%).In 6 eyes (26.1%), only a few hyperautofluorescent spots scattered in the macula were observed.SD-OCT demonstrated significantly reduced (by 69.5% on average) or even disappearance of subretinal fluid in the eyes.The fluorescence intensity in FAF imaging showed a significant positive correlation with the volume of subretinal fluid detected by SD-OCT (r=0.626, P < 0.05). CONCLUSIONS: The combination of fluorescein angiography, FAF imaging and SD-OCT can significantly improve the diagnostic accuracy of VKH disease.FAF imaging combined with SD-OCT provides an effective and noninvasive modality for evaluation of remission and monitoring the changes in VKH disease.

Chronic Neuroinflammation Induced by Lipopolysaccharide Injection into the Third Ventricle Induces Behavioral Changes.

The existence of Gram-negative bacteria in the brain, regardless of underlying immune status has been demonstrated by recent studies. The colocalization of lipopolysaccharide (LPS) with Aß1-40/42 in amyloid plaques supports the hypothesis that brain microbes may be the cause, triggering chronic neuroinflammation, leading to Alzheimer's disease (AD). To investigate the behavioral changes induced by infectious neuroinflammation, we chose the third ventricle as the site of a single LPS injection (20 µg or 80 µg) in male Wistar rats to avoid mechanical injury to forebrain structures while inducing widespread inflammation throughout the brain. Chronic neuroinflammation induced by LPS resulted in depressive-like behaviors and the impairment of spatial learning; however, there was no evidence of the development of pathological hallmarks (e.g., the phosphorylation of tau) for 10 months following LPS injection. The acceleration of cholesterol metabolism via CYP46A1 and the retardation of cholesterol synthesis via HMGCR were observed in the hippocampus of rats treated with either low-dose or high-dose LPS. The rate-limiting enzymes of cholesterol metabolism (CYP46A1) in SH-SY5Y cells and synthesis (HMGCR) in U251 cells were altered by inflammation stimulators, including LPS, IL-1ß, and TNF-α, through the TLR4/MyD88/NF-κB signaling pathway. The data suggest that chronic neuroinflammation provoked by the administration of LPS into the third ventricle may induce depressive-like symptoms and that the loss of cholesterol might be a biomarker of chronic neuroinflammation. The lack of pathological hallmarks of AD in our model indicates that Gram-negative bacteria infection might not be a single cause of AD.

Development and external validation of a COVID-19 mortality risk prediction algorithm: a multicentre retrospective cohort study.

OBJECTIVE: This study aimed to develop and externally validate a COVID-19 mortality risk prediction algorithm. DESIGN: Retrospective cohort study. SETTING: Five designated tertiary hospitals for COVID-19 in Hubei province, China. PARTICIPANTS: We routinely collected medical data of 1364 confirmed adult patients with COVID-19 between 8 January and 19 March 2020. Among them, 1088 patients from two designated hospitals in Wuhan were used to develop the prognostic model, and 276 patients from three hospitals outside Wuhan were used for external validation. All patients were followed up for a maximal of 60 days after the diagnosis of COVID-19. METHODS: The model discrimination was assessed by the area under the receiver operating characteristic curve (AUC) and Somers' D test, and calibration was examined by the calibration plot. Decision curve analysis was conducted. MAIN OUTCOME MEASURES: The primary outcome was all-cause mortality within 60 days after the diagnosis of COVID-19. RESULTS: The full model included seven predictors of age, respiratory failure, white cell count, lymphocytes, platelets, D-dimer and lactate dehydrogenase. The simple model contained five indicators of age, respiratory failure, coronary heart disease, renal failure and heart failure. After cross-validation, the AUC statistics based on derivation cohort were 0.96 (95% CI, 0.96 to 0.97) for the full model and 0.92 (95% CI, 0.89 to 0.95) for the simple model. The AUC statistics based on the external validation cohort were 0.97 (95% CI, 0.96 to 0.98) for the full model and 0.88 (95% CI, 0.80 to 0.96) for the simple model. Good calibration accuracy of these two models was found in the derivation and validation cohort. CONCLUSION: The prediction models showed good model performance in identifying patients with COVID-19 with a high risk of death in 60 days. It may be useful for acute risk classification. WEB CALCULATOR: We provided a freely accessible web calculator (https://www.whuyijia.com/).

[Construction of hp0523 gene mutant in Helicobacter pylori Cag-PAI and the influence on the ability of CagA protein translocation].

OBJECTIVE: To construct the hp0523 gene mutant of Helicobacter pylori and investigate the function of hp0523 gene. METHODS: We designed and amplified the upstream homologous fragment and downstream homologous fragment of hp0523 gene via PCR method. We constructed the suicide plasmid pBlueKM40-deltahp0523 based on allelic exchange. We introduced the suicide plasmid pBlueKM40-deltahp0523 into Helicobacter pylori 11637 by electroporation and screened the mutant based on antibiotic selection. We checked the mutant using the PCR and gene sequenced. We performed the coculture of Helicobacter pylori and gastric cell BGC-823 and detected the ability of CagA's translocation and expression via Western blot. RESULTS: We constructed the suicide plasmid pBlueKM40-deltahp0523 successfully and got the hp0523 deletion mutant. PCR and gene sequenced results showed the gene hp0523 was deleted. The results of CagA translocation assay showed that hp0523 interrupted the translocation of CagA. The comparison between wild-type and mutant showed that hp0523 affected the expression of CagA. CONCLUSIONS: We constructed the hp0523 deletion mutant of Helicobacter pylori NCTC11637. This study suggests that hp0523 gene is an important virulence factor, which may be a component of the apparatus for CagA's translocation.

Dynamic Analysis of Perioperative Hidden Blood Loss in Intertrochanteric Fractures.

To analyze the dynamic variation in perioperative hidden blood loss in patients with intertrochanteric fracture. From January to December 2017, 79 patients with intertrochanteric fracture were treated with proximal femoral nail antirotation. Serial complete blood count assays were performed consecutively in the 3 days after admission, on the day of surgery, and 7 days postoperatively. Blood loss during surgery, postoperative drainage, and perioperative blood transfusion volumes were recorded. Dynamic changes in hemoglobin (Hb) prior to surgery were recorded and compared between males and females. Patients were divided into the no blood transfusion group, the 400-mL blood transfusion group, and the 800-mL blood transfusion group depending on the volume of perioperative blood transfusion. Total and hidden blood loss were separately calculated according to the Gross equation. Lowest mean Hb values occurred on day 2 after admission among men (104.8 g/L) and on day 3 after admission among women (98.6 g/L). The average Hb decrease was 11.4 g/L, 11.8 g/L, and 8.9 g/L in the no, 400-mL, and 800-mL blood transfusion groups, respectively. The lowest Hb value occurred on postoperative day 2. Hemoglobin increased on postoperative day 3 and stabilized by day 6. In the no blood transfusion group, the average total blood loss was 406.0 ± 255.6 mL, 628.3 ± 267.2 mL, and 759.7 ± 322.1 mL in the no blood transfusion, 400-mL blood transfusion, and 800-mL blood transfusion groups, respectively, and hidden blood loss was 326.0 ± 246.6 mL, 512.1 ± 247.3 mL, and 596.1 ± 306.9 mL, respectively. Perioperative hidden blood loss occurred prior to surgery for intertrochanteric fracture and ended on postoperative day 2.

Step-advanced rectangular flap: A novel technique for the reconstruction of soft-tissue defects overlying the Achilles tendon in children (an observational study).

Soft-tissue defects overlying the Achilles tendon are common complications after bicycle or motorcycle spoke injuries in children and usually require surgical management by various flaps. There is no apparent consensus on the optimal choice of flaps for these injuries. We designed a novel step-advanced rectangular flap to reconstruct small to moderate soft-tissue defects around the Achilles tendon. This study was performed to review our experience and evaluate the clinical effectiveness of the step-advanced rectangular flap.From May, 2014 to September, 2016, 12 consecutive children with soft-tissue defects overlying the Achilles tendon caused by spoke injuries were treated with the step-advanced rectangular flap. The patients' general information, surgical details, and postoperative complications were recorded. The Mazur evaluation system was used to assess clinical outcomes.All patients were followed up for ≥12 months (range 12-38 months). All flaps survived completely. Superficial infection occurred in 2 patients and healed by second intention after dressing changes; the other patients' surgical wounds healed by primary intention. The scars around the flaps in 2 patients were remarkable, and all others showed good results in terms of flap color and texture. Ankle function was normal, and satisfactory results were obtained in all cases. According to the Mazur evaluation system, the results were excellent in 9 patients and good in 3, with an excellent and good rate of 100% at 12 months postoperatively.The rectangular advancement flap appears to be a simple and reliable method for small to moderate soft tissue defects overlying the Achilles tendon in children.

The effect of tranexamic acid on hidden bleeding in older intertrochanteric fracture patients treated with PFNA.

OBJECTIVE: To investigate the effect of tranexamic acid (TXA) on hidden bleeding in older intertrochanteric fracture patients treated with intramedullary nails. METHOD: Between January 2016 and January 2017, 100 cases of intertrochanteric fractures eligible for the study were treated with proximal femoral nail antirotation (PFNA) in our hospital. All patients were divided into two groups of 50 patients each: the TXA group and a blank control group. In the TXA group, all patients received TXA at a dose of 10 mg/kg-1 intravenously, 10 min preoperatively and 5 h postoperatively. The control group did not receive TXA. We recorded the volume of intraoperative blood loss and postoperative drainage, and the need for postoperative blood transfusion and transfusion volume for all patients. Blood routine examination was performed on the day of surgery and 2 days postoperatively. We calculated the total blood loss and hidden blood loss in the two groups separately according to the Gross equation. All patients underwent deep vein ultrasound of the lower limbs preoperatively and 1 week postoperatively to detect thrombosis. RESULTS: Compared with controls, patients in the TXA group had lower: overt bleeding (50.59 ml; p = .012), total blood loss (181.58 ml; p = .005), hidden blood loss (130.64 ml; p = .037), volume of blood transfusion (110.0 ml; p = .019), and 20% lower transfusion rate compared with the control group. Patients receiving short-nail fixation had significantly lower hidden blood loss compared with patients receiving long-nail fixation (p < .05). However, we found no statistically significant difference in the incidence of deep vein thrombosis in the lower limbs between the two groups (p = .938). CONCLUSION: TXA significantly reduced hidden blood loss in older intertrochanteric fracture patients treated with intramedullary nails without an increased risk of thrombosis in lower limb deep veins.

Pigment epithelium-derived factor protects retinal ganglion cells from hypoxia-induced apoptosis by preventing mitochondrial dysfunction.

AIM: To investigate the potential of pigment epithelium-derived factor (PEDF) to protect the immortalized rat retinal ganglion cells-5 (RGC-5) exposed to CoCl2-induced chemical hypoxia. METHODS: After being differentiated with staurosporine (SS), RGC-5 cells were cultured in four conditions: control group cells cultured in Dulbecco's modified eagle medium (DMEM) supplemented with 10% fetal bovine serum, 100 µmol/mL streptomycin and penicillin (named as normal conditions); hypoxia group cells cultured in DMEM containing 300 µmol/mL CoCl2; cells in the group protected by PEDF were first pretreated with 100 ng/mL PEDF for 2h and then cultured in the same condition as hypoxia group cells; and PEDF group cells that were cultured in the presence of 100 ng/mL PEDF under normal conditions. The cell viability was assessed by MTT assay, the percentage of apoptotic cells was quantified using Annexin V-FITC apoptosis kit, and intra-cellar reactive oxygen species (ROS) was measured by dichloro-dihydro-fluorescein diacetate (DCFH-DA) probe. The mitochondria-mediated apoptosis was also examined to further study the underlying mechanism of the protective effect of PEDF. The opening of mitochondrial permeability transition pores (mPTPs) and membrane potential (Δψm) were tested as cellular adenosine triphosphate (ATP) level and glutathione (GSH). Also, the expression and distribution of Cyt C and apoptosis inducing factor (AIF) were observed. RESULTS: SS induced differentiation of RGC-5 cells resulting in elongation of their neurites and establishing contacts between outgrowths. Exposure to 300 µmol/mL CoCl2 triggered death of 30% of the total cells in cultures within 24h. At the same time, pretreatment with 100 ng/mL PEDF significantly suppressed the cell death induced by hypoxia (P<0.05). The apoptosis induced by treatment of CoCl2 was that induced cell death accompanied with increasing intra-cellar ROS and decreasing GSH and ATP level. PEDF pre-treatment suppressed these effects (P<0.05). Additionally, PEDF treatment inhibited the opening of mPTPs and suppressed decreasing of Δψm in RGC-5 cells, resulting in blocking of the mitochondrial apoptotic pathway. CONCLUSION: Pretreatment of RGC-5 cells with 100 ng/mL PEDF significantly decreases the extent of apoptosis. PEDF inhibits the opening of mPTPs and suppresses decreasing of Δψm. Moreover, PEDF also reduces ROS production and inhibits cellular ATP level's reduction. Cyt C and AIF activation in PEDF-pretreated cultures are also reduced. These results demonstrate the potential for PEDF to protect RGCs against hypoxic damage in vitro by preventing mitochondrial dysfunction.