Pseudomonas aeruginosa colonization enhances ventilator-associated pneumonia-induced lung injury.

BACKGROUND: Pseudomonas aeruginosa (PA) is the single-most common pathogen of ventilator-associated pneumonia (VAP). Large quantities of PA in the trachea of ventilated patients are associated with an increased risk of death. However, the role of PA colonization in PA VAP-induced lung injury remains elusive. This study examined the effect and mechanism of PA colonization in VAP-induced lung injury. METHODS: C57BL/6 wild-type (WT) and c-Jun N-terminal kinase knockout (JNK1(-/-)) mice received mechanical ventilation for 3 h at 2 days after receiving nasal instillation of PA (1 × 10(6) colony forming unit) or normal saline. RESULTS: Intranasal instillation of PA or mechanical ventilation induced the expression of interleukin-6 (IL-6) in the lungs. Phospho-JNK protein expression in the lungs was significantly increased in mice receiving mechanical ventilation after PA instillation as compared with those receiving ventilation alone. Mechanical ventilation after PA instillation significantly increased the expression of tumor necrosis factor-α (TNF-α), IL-1ß, and macrophage inflammatory protein-2 (MIP-2) proteins; neutrophil sequestration; and TNF-α, IL-1ß, and IL-6 levels in the lungs of WT mice, but not in JNK1(-/-) mice. CONCLUSION: PA colonization plays an important role in PA VAP-induced lung injury through the induction of JNK1-mediated inflammation. PA-induced VAP causes lung injury through JNK signaling pathway in the lungs. JNK inhibition in ICU patients with higher percentages of PA colonization may reduce VAP-induced lung injury and mortality.

Outcomes of cervical degenerative disc disease treated by anterior cervical discectomy and fusion with self-locking fusion cage.

BACKGROUND: Cervical degenerative disc (CDD) disease is a common type of spondylosis. Although anterior cervical discectomy and fusion (ACDF) is the preferred treatment for CDD disease, internal fixation with a titanium plate may cause various complications. The invention of the ACDF with a self-locking fusion cage (ROI-C) has effectively decreased the incidence of postoperative complications. AIM: To observe the outcomes of CDD disease treated by ACDF with a ROI-C. METHODS: Ninety patients with CDD disease treated at our hospital from March 2019 to March 2021 were included. They were divided into two groups (control group and observation group, n = 45 in each) using a random number table. Patients in the control group received ACDF plus internal fixation with a titanium plate. Those in the observation group received ACDF + ROI-C placement. The two groups of patients were compared in terms of surgical parameters, pain, cervical spine function, range of motion, and complications. RESULTS: The two groups of patients showed no significant differences in surgical time, blood loss, drainage volume, and length of hospital stay (P > 0.05). No significant differences in the visual analogue scale (VAS), Japanese Orthopedic Association (JOA), and neck disability index (NDI) scores were observed between the two groups before surgery (P > 0.05). The VAS and NDI scores in the observation group were considerably lower than those in the control group after surgery; however, the JOA scores in the observation group were significantly higher than those in the control group (P < 0.05). No significant differences were observed in cervical disc height and the range of motion of the superior or inferior adjacent vertebrae between the two groups before surgery (P > 0.05). The disc height in the observation group was larger than that in the control group after surgery. The range of motion of both the superior and inferior adjacent vertebrae was significantly smaller in the observation group than in the control group (P < 0.05). The incidence of complications was only 2.22% in the observation group compared to 15.56% in the control group, and the difference was statistically significant (P < 0.05). CONCLUSION: Cervical spine function restoration was better with ROI-C with internal fixation in ACDF than with conventional titanium plates in ACDF for CDD disease.

Effectiveness and postoperative rehabilitation of one-stage combined anterior-posterior surgery for severe thoracolumbar fractures with spinal cord injury.

BACKGROUND: Thoracolumbar fractures are generally combined with spinal cord injury to varying degrees, which may cause deterioration of the patients' condition and increase the difficulty of clinical treatment. At present, anterior or combined anterior-posterior surgery is preferred for severe thoracolumbar fractures. AIM: To investigate the effectiveness and postoperative rehabilitation of one-stage combined anterior-posterior surgery for severe thoracolumbar fractures with spinal cord injury. METHODS: One-hundred-and-twenty patients who received surgery for severe thoracolumbar fractures with spinal cord injury at our hospital from February 2018 to February 2020 were randomly enrolled. They were randomly divided into group 1 (one-stage combined anterior-posterior surgery, n = 60) and group 2 (one-stage anterior-approach surgery, n = 60). Treatment efficacy was compared between the two groups. RESULTS: Blood loss was greater and the operation time was longer in group 1 than in group 2, and the differences were statistically significant (P < 0.05). Incision length, intraoperative X-rays, and length of hospital stay were not significantly different between the two groups (P > 0.05). Preoperative function of the affected vertebrae was not significantly different between the two groups (P > 0.05). In each group, the patients showed significant improvement after surgery. The anterior vertebral height ratio and the posterior vertebral height ratio in group 1 after surgery were significantly higher than those in group 2. The Cobb angle after surgery was significantly lower in group 1 than in group 2 (P < 0.05). The canal-occupying ratio of the affected vertebrae was not significantly different between the two groups (P > 0.05). Before surgery, there was no significant difference in the quality of life scores between the two groups (P > 0.05). The above indicators were significantly improved after surgery compared with before surgery in each group. In addition, these indicators were markedly better in group 1 than in group 2 after surgery (P < 0.05 for each). CONCLUSION: One-stage combined anterior-posterior surgery effectively improves the function of the affected vertebrae and the life quality of patients with severe thoracolumbar fractures and spinal cord injury. This surgical approach is worthy of popularization in clinical use.

Tear cytokine profile in medicated glaucoma patients: effect of monocyte chemoattractant protein 1 on early posttrabeculectomy outcome.

PURPOSE: To determine the tear cytokine profile from medicated glaucoma patients scheduled for trabeculectomy and to establish whether a specifically elevated cytokine level is related to early postoperative scarring. DESIGN: Prospective case-control study. PARTICIPANTS: Sixty-one patients treated with topical antiglaucoma medications and 29 normal subjects with no prior topical treatment were recruited for the study. METHODS: Schirmer strips were used to collect tear samples. A multiplex bead assay was used to quantify the presence of proinflammatory cytokines in the tears. The patients were followed up for 6 months after surgery to determine whether any postoperative intervention to maintain filtering bleb function was required. MAIN OUTCOME MEASURES: The level of cytokines in tear specimens from medicated glaucoma patients was the main outcome measure for the study. The need for postoperative bleb needling within 6 months was a secondary outcome measure. RESULTS: Of the 17 cytokines assayed, only monocyte chemoattractant protein 1 (MCP-1) was elevated significantly in the medicated eyes compared with the unmedicated eyes (P < 0.0004). At 6 months after surgery, 18 (30%) of the 61 eyes required postoperative intervention. A much higher MCP-1 level was detected in these eyes compared with the remaining 43 that did not require intervention (P < 0.0001). The duration of use of topical medication correlated with increasing levels of MCP-1, although the types of glaucoma medication and the number of bottles of medications did not have any significant relationship with the level of MCP-1. CONCLUSIONS: In tears from topically medicated glaucoma eyes in an Asian population, MCP-1 was found to be the predominant cytokine elevated. Eyes with a propensity to scar in the early postoperative period have a significantly raised level of MCP-1. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found after the references.

A novel Pro126His beta propeller mutation in integrin alphaIIb causes Glanzmann thrombasthenia by impairing progression of pro-alphaIIbbeta3 from endoplasmic reticulum to Golgi.

BACKGROUND: Glanzmann thrombasthenia (GT) is an autosomal recessive bleeding disorder characterized by lack of platelet aggregation in response to most physiological agonists and caused by either a lack or dysfunction of the platelet integrin alphaIIbbeta3 (glycoprotein IIb/IIIa). PATIENTS: Mucocutaneous bleeding manifestations and platelet dysfunction consistent with GT were observed in a 20-year-old proband of a Chinese family. OBJECTIVES: To determine the molecular basis of GT and characterize the mutation by in vitro expression studies. RESULTS: Analysis of the patient's platelets by fluorescence-activated cell sorting demonstrated the presence of trace amounts of beta3, exposed on her platelet surface, but a complete absence of alphaIIbbeta3. Sequence analysis revealed a novel C470A transversion in exon 4 of the alphaIIb gene predicting a Pro126His alteration in the blade 2 of the alphaIIb beta propeller domain. The proband was homozygous for the mutation, the mother and the father were heterozygous, whereas 100 healthy subjects lacked this transversion. Chinese hamster ovary cells cotransfected with cDNAs of mutated alphaIIb and wild-type beta3 failed to express alphaIIbbeta3 on the cell surface as shown by FACS. Western blot analysis of the cell lysates showed no detectable mature alphaIIb. Immunoprecipitation with antibody against beta3 demonstrated pro-alphaIIb in the cells expressing the mutant alphaIIbbeta3, indicating pro-alphaIIbbeta3 complex formation. Intracellular immunofluorescence studies demonstrated the pro-alphaIIbbeta3 complex that co-localized with an ER marker, but showed minimal co-localization with a Golgi marker. CONCLUSIONS: A novel Pro126His mutation in alphaIIb compromised transport of the pro-alphaIIbbeta3 complex from the endoplasmic reticulum to the Golgi, leading to intracellular retention. The impaired alphaIIbbeta3 transport is responsible for the thrombasthenia in this patient.

[Molecular mechanisms of Glanzmann thrombasthenia caused by alphaII b P126H mutation: an in vitro experiment].

OBJECTIVE: To explore the molecular mechanisms of Glanzmann thrombasthenia caused by alpha IIb P126H mutation. METHODS: Eukaryotic vector of alpha IIb P126H was constructed by PCR site-directed mutagenesis and then co-transfected with eukaryotic vector PCDM8 II a expressing the subunit beta3 into human renal epithelial cells of the line 293& and Chinese hamster ovarian cancer cells of the line CHO after sequencing. The membrane expression of alpha IIb P126H mutant was analyzed by flow cytometry and the whole expression was confirmed by Western blotting. The alpha IIb P126H mutant subcellular localization was determined by laser confocal scanning microscopy. RESULTS: The 293T cells cotransfected with cDNAs of mutated alpha IIb and wild-type beta3 failed to express alpha II bbeta3 on the cell surface as shown by FACS. Western blotting of the cell lysate showed no detectable mature alpha IIb. Immunofluorescence studies demonstrated proa II bbeta3 complex colocalized with an endoplasmic reticulum (ER) marker, but showed minimal colocalization with an Golgi marker. CONCLUSION: The P126H mutation in alpha IIb prevents transport of the pro-alpha II bbeta3 complex from ER to the Golgi body, thus hindering its maturation and surface expression. The impaired alpha II bp33 transport is responsible for the thrombasthenia.

Pseudomonas aeruginosa Ventilator-Associated Pneumonia Induces Lung Injury through TNF-α/c-Jun NH2-Terminal Kinase Pathways.

Ventilator-associated pneumonia (VAP) is a common nosocomial infection among intensive care unit (ICU) patients. Pseudomonas aeruginosa (PA) is the most common multidrug-resistant Gram-negative pathogen and VAP caused by PA carries a high rate of morbidity and mortality. This study examined the molecular mechanism of PA VAP-induced lung injury. C57BL/6 wild-type (WT) mice and JNK1 knockout (JNK1-/-) mice received mechanical ventilation (MV) for 3 h at 2 days after receiving nasal instillation of PA. The WT and JNK1-/- mice also received MV after the induction of lung injury by instillation of supernatants from PA-stimulated alveolar macrophages (AMs). AMs isolated from WT, IκB-kinase (IKK)ßΔMye (IKKß was selectively deleted in macrophages), and JNK1-/- mice were ex vivo stimulated with live PA and supernatants were collected for cytokine assay. Intranasal instillation of 106 PA enhanced MV-induced NF-κB DNA binding activity in the lungs and nitrite levels in BALF. MV after PA instillation significantly increased the expression of ICAM and VCAM in the lungs and TNF-α, IL-1ß, and IL-6 levels in bronchoalveolar lavage fluid (BALF) of WT mice, but not in JNK1-/- mice. MV after supernatant instillation induced more total protein concentration in BALF and neutrophil sequestration in the lungs in WT mice than JNK1-/- mice and cytokine assay of supernatants indicated that TNF-α is a critical regulator of PA VAP-induced lung injury. Ex vivo PA stimulation induced TNF-α production by AMs from WT as well as JNK1-/- mice but not IKKßΔMye mice. In summary, PA colonization plays an important role in PA VAP-induced lung injury through the induction of JNK1-mediated inflammation. These results suggest that the pathogenesis mechanism of PA VAP involves production of TNF-α through activation of IKK/NF-κB pathways in AMs and JNK signaling pathway in the lungs.

Lauric Acid Is an Inhibitor of Clostridium difficile Growth in Vitro and Reduces Inflammation in a Mouse Infection Model.

Clostridium difficile is a Gram-positive, spore-forming anaerobic human gastrointestinal pathogen. C. difficile infection (CDI) is a major health concern worldwide, with symptoms ranging from diarrhea to pseudomembranous colitis, toxic megacolon, sepsis, and death. CDI onset and progression are mostly caused by intestinal dysbiosis and exposure to C. difficile spores. Current treatment strategies include antibiotics; however, antibiotic use is often associated with high recurrence rates and an increased risk of antibiotic resistance. Medium-chain fatty acids (MCFAs) have been revealed to inhibit the growth of multiple human bacterial pathogens. Components of coconut oil, which include lauric acid, have been revealed to inhibit C. difficile growth in vitro. In this study, we demonstrated that lauric acid exhibits potent antimicrobial activities against multiple toxigenic C. difficile isolates in vitro. The inhibitory effect of lauric acid is partly due to reactive oxygen species (ROS) generation and cell membrane damage. The administration of lauric acid considerably reduced biofilm formation and preformed biofilms in a dose-dependent manner. Importantly, in a mouse infection model, lauric acid pretreatment reduced CDI symptoms and proinflammatory cytokine production. Our combined results suggest that the naturally occurring MCFA lauric acid is a novel C. difficile inhibitor and is useful in the development of an alternative or adjunctive treatment for CDI.

[A preliminary report on bimanual microphacoemulsification].

OBJECTIVE: To evaluate the surgical technique, feasibility, and outcome of bimanual microphacoemulsification. METHODS: The preliminary clinical study included 132 senile cataract eyes. A temporal clear cornea incision was made using 19G microvitreoretinal blade with the exterior incision length of 1.4 mm, the interior incision length of 1.2 mm, and the tunnel length of 1.0 mm. A 1.2 mm x 1.0 mm clear cornea side port was created with 19G microvitreoretinal blade at 12 o'clock in the right eye or 6 o'clock in the left eye. A sleeveless titanium phaco needle with an outer diameter of 0.9 mm was inserted through the temporal clear cornea incision. An irrigating chopper was inserted through the side port as the left-hand instrument, bimanual nucleofractis and nuclear emulsification were performed using quick chop technique. The lens cortical removal was performed bimanually with the Duet Bimanual I/A System. The study parameters included phacoemulsification time, intraoperative complications and early postoperative outcome. RESULTS: The mean Phacoemulsification time was 0.75 +/- 0.64 min. vision acuity equal or better than 0.5 were 55.30%, 87.12% and 90.15% after surgery one day, one week and one mother respectively. Eyes with 0.5 best corrected visual acuity amounted to 90.91%, while with 0.8, amounted to 77.27% all the treated eyes 1 month postoperation. The formation of anterior chamber was successfully maintained in every case. There was various degree of nuclear hardness in all of case studied. CONCLUSION: Bimanual microphacoemulsification is a feasible, secure, and effective surgery for cataract extraction through a sub-1.5 mm clear cornea incision.

Changes in chloroplast ultrastructure, fatty acid components of thylakoid membrane and chlorophyll a fluorescence transient in flag leaves of a super-high-yield hybrid rice and its parents during the reproductive stage.

In plants, it is well established that chloroplast is one of the early targgeted organelles to breakdown during leaves senescing. Here we applied a newly developed super-high-yield hybrid rice (Oryza sativa) LiangYouPeiJiu (LYPJ) and its parents lines to investigate changes in ultrastructure of chloroplasts, fatty acid composition of thylakoid membrane lipids and chlorophyll (Chl) a fluorescence transient in natural senescing leaves. We found that at full expansion of flag leaves in three lines, chloroplasts often showed oblong shapes with a typical membrane system of stroma and grana thylakoids, whereas their shapes had been changed from oblong to spherical during senescence. Our data showed that the initiation of senescence displayed accumulation of starch and an increase in the number and size of plastoglobuli with the damaged thylakoid membranes; subsequently, swollen thylakoid membranes in stroma and in grana with a significant increase in MDA content, and disorganization of thylakoid membrane system with significant changes in fatty acid composition of thylakoid membrane lipids were developed. Compared with its parents, the newly developed hybrid rice LYPJ had the highest capacity of carbohydrate transport from leaves (sources) to ears (sink), marked with the lowest accumulation of starch grains in the leaf chloroplasts, and the slowest senescing rate of chloroplast in overall leaf senescence process. Chl a fluorescence transients of three kinds of flag leaves were analyzed by so-called JIP-test. The results of analysis suggest that these findings inculding a high inherited activity of antioxidant enzymes and high photosynthetate transport to pretect chloroplast structure in the hybrid rice LYPJ have close relations to its super-high yield.

[Molecular mechanisms of Glanzmann thrombasthenia caused by alpha II b L721R and Q860X compound heterozygous mutation].

OBJECTIVE: To explore the molecular mechanisms of Glanzmann thrombasthenia caused by alpha II b L721R and Q860X compound heterozygous mutation. METHODS: All exons and exon-intron boundaries of alpha II b and beta3 gene were amplified by PCR and analyzed by direct DNA sequencing. Gene polymorphisms were excluded by direct DNA sequencing. Alpha II b L721R and Q860X mutants expressing vectors were constructed by in vitro site-directed mutagenesis. The expression of alpha II b L721R and Q860X mutants on transfected cell membrane were analyzed by flow cytometry and the whole expression level was confirmed by Western blot. The subcellular localizations of alpha II b L721R and Q860X mutants were determined by immunofluorescent confocal scanning microscopy. RESULTS: The alpha II b compound heterozygous mutations, T2255G (L721R) and C2671T (Q860X), were identified in the proband, the former being inherited from the maternal side and the latter the paternal side. The 293T cells cotransfected with mutated alpha II b L721R and wild-type beta3 expression plasmids expressed 2.1% of normal amount of alpha II b on the cell surface as shown by FACS, in contrast to 31.9% of normal amount of alpha II b on the cells cotransfected with cDNAs of mutated alpha II b Q860X and wildtype beta3 expression plasmids. Western blot of the cell lysates showed no detectable mature alpha II b in cells lysates with L721R mutant. While, truncated alpha II b protein was detected in cell lystes with Q860X mutant. Immunofluorescence studies demonstrated that both L721R and Q860X mutant pro-alpha II bbeta33 complex colocalized in endoplasmic reticulum, but a little in Golgi. CONCLUSIONS: The L721R and Q860X mutations of alpha II b prevent transport of the pro-alpha II bbeta3 complex from the endoplasmic reticulum to the Golgi, hindering its maturation and surface expression. The impaired alpha II bbeta3 transport is responsible for the thrombasthenia.