[Clinical observation on the treatment of ossification of the posterior longitudinal ligament of the cervical spine using 3D printed self-stable zero-profile artificial vertebral body].

Objective: To observe the clinical efficacy of a 3D printed self-stable zero-profile artificial vertebral body for anterior cervical corpectomy decompression and fusion in the treatment of ossification of the posterior longitudinal ligament (OPLL) of the cervical spine. Methods: In this prospective randomized controlled trial, patients diagnosed with OPLL in Luohe Central Hospital from January to July 2022 were divided into a zero-profile group (3D printed self-stable zero-profile artificial vertebral body was used for internal fixation and fusion after anterior cervical subtotal decompression,) and titanium-mesh group (titanium-mesh and titanium plate were used for internal fixation and fusion after anterior cervical subtotal decompression) according to envelope random method. Operation time, intraoperative blood loss, Japanese Orthopaedic Association (JOA) score and improvement rate, incidence of postoperative prosthesis subsidence, and bone graft fusion were recorded and compared between the two groups. Results: Finally, 21 patients in the zero notch group and 20 patients in the titanium mesh group were included in the study and were followed-up. In the zero-profile group, there were 16 males and 5 females, aged (48.0±12.7) years. In the titanium-mesh group, there were 14 males and 6 females, aged (49.8±10.2) years. All the 41 patients successfully completed the operation. In the zero-profile group, the surgical time was (50.04±8.45) minutes, the blood loss was (95.38±26.07) ml and the hospitalization cost was (42.32±6.12) thousand yuan. In the titanium-mesh group, the surgical time was (59.20±11.95) minutes, the blood loss was (93.10±27.86) ml and the hospitalization cost was (42.10±6.71) thousand yuan. The surgical time in the zero-profile group was shorter than that in the titanium-mesh group (P=0.007), and there was no statistically significant difference in blood loss and hospitalization costs between the two groups (both P>0.05). The 41 patients were followed-up for (14.29±1.45) months. Four cases (20.0%) in the titanium mesh group experienced swallowing difficulties and 0 cases in the zero incision group, the difference between the two groups was statistically significant (P=0.048). No intraoperative hematoma, spinal cord nerve recompression, airway crisis, incision infection complications, and no steel plate or screw breakage or displacement occurred after surgery. At the last follow-up, all cases had bone fusion. At the follow-up of 12 months after surgery, the JOA score of the zero incision group increased from preoperative (10.33±1.71) points to (15.47±0.81) points, with an improvement rate of 76.1%±15.7%; the JOA score of the titanium mesh group increased from (10.30±1.75) points to (15.30±0.92) points, with an improvement rate of 73.2%±16.7%; there was no statistically significant difference in improvement rate between the two groups (P=0.580). At the follow-up of 12 months after surgery, 1 case (4.8%) in the zero incision group and 8 cases (40.0%) in the titanium mesh group experienced implant sinking, and the difference between the two groups was statistically significant (P=0.009). Conclusion: Compared with titanium-mesh, 3D printed self-stable zero-profile artificial vertebral body for the treatment of OPLL of the cervical spine can achieve good surgical efficacy, shorter surgical time, lower incidence of postoperative chronic swallowing discomfort, and can provide a better bone material bonding interface and be less prone to prosthesis settlement.

[Analysis of iodine nutritional status of children aged 8-10 years in Zhejiang Province from 2016 to 2021].

Objective: To analyze the iodine nutrition status of children aged 8 to 10 years in Zhejiang Province from 2016 to 2021. Methods: A multi-stage stratified sampling method was used to select non-residential children aged 8 to 10 years from 90 counties in Zhejiang Province. A total of 114 103 children were included in the study from 2016 to 2021. Direct titration method and arsenic-cerium catalytic spectrophotometry were used to detect salt iodine content and urinary iodine level, respectively, to evaluate the iodine nutritional status of children. Ultrasound was used to detect thyroid volume and analyze the current prevalence of goiter in school-age children. Results: The age of 114 103 children was (9.04 ± 0.81) years old, with 50.0% of (57 083) boys. The median of iodine content M (Q1, Q3) in children's household salt was 23.00 (19.80, 25.20) mg/kg, including 17 242 non-iodized salt, 6 173 unqualified iodized salt, and 90 688 qualified iodized salt. The coverage rate of iodized salt was 84.89%, and the coverage rate of qualified iodized salt was 79.48%. The proportion of non-iodized salt increased from 11.85% in 2016 to 16.04% in 2021 (χ2trend=111.427, P<0.001). The median of urinary iodine concentration M (Q1, Q3) in children was 182.50 (121.00, 261.00) µg/L, among which the proportions of iodine deficiency, iodine suitability, iodine over suitability, and iodine excess were 17.25% (19 686 cases), 39.21% (44 745 cases), 26.85% (30 638 cases), and 16.68% (19 034 cases), respectively. The median of urinary iodine concentration in children in inland areas [M (Q1, Q3): 190.90 (128.80, 269.00) µg/L] was significantly higher than that in children in coastal areas [M (Q1, Q3): 173.00 (113.00, 250.30) µg/L] (P<0.001). From 2016 to 2021, a total of 39 134 ultrasound examinations were conducted, and 1 229 cases of thyroid enlargement were detected. The goiter rate was 3.14% (95%CI: 2.97%-3.32%). The incidence of goiter in children in coastal areas [3.45% (95%CI: 3.19%-3.72%), 641/18 604] was higher than that in children in inland areas [2.86% (95%CI: 2.64%-3.10%), 588/20 530] (P=0.001). Conclusion: From 2016 to 2021, the iodine nutrition level of children aged 8-10 years in Zhejiang Province is generally suitable, and the rate of goiter in children meets the limit of iodine deficiency disease elimination standards.

[A phase I study of subcutaneous envafolimab (KN035) monotherapy in Chinese patients with advanced solid tumors].

Objective: To evaluate the safety and antitumor activity of envafolimab monotherapy in Chinese patients with advanced solid tumors. Methods: This open-label, multicenter phase I trial included dose escalation and dose expansion phases. In the dose escalation phase, patients received subcutaneous 0.1, 0.3, 1.0, 2.5, 5.0 or 10.0 mg/kg envafolimab once weekly (QW) following a modified "3+ 3" design. The dose expansion phase was performed in the 2.5 mg/kg and 5.0 mg/kg (QW) dose cohorts. Results: At November 25, 2019, a total of 287 patients received envafolimab treatment. During the dose escalation phase, no dose-limiting toxicities (DLT) was observed. In all dose cohorts, drug-related treatment-emergent adverse events (TEAEs) for all grades occurred in 75.3% of patients, and grade 3 or 4 occurred in 20.6% of patients. The incidence of immune-related adverse reactions (irAE) was 24.0% for all grades, the most common irAEs (≥2%) included hypothyroidism, hyperthyroidism, immune-associated hepatitis and rash. The incidence of injection site reactions was low (3.8%), all of which were grades 1-2. Among the 216 efficacy evaluable patients, the objective response rate (ORR) and disease control rate (DCR) were 11.6% and 43.1%, respectively. Median duration of response was 49.1 weeks (95% CI: 24.0, 49.3). Pharmacokinetic (PK) exposure to envafolimab is proportional to dose and median time to maximum plasma concentration is 72-120 hours based on the PK results from the dose escalation phase of the study. Conclusion: Subcutaneous envafolimab has a favorable safety and promising preliminary anti-tumor activity in Chinese patients with advanced solid tumors.

[A prospective cohort study of long-term fasting blood glucose variability and risk of mortality in patients with type 2 diabetes].

Objective: To investigate the association between long-term fasting blood glucose (FPG) variability and all-cause mortality in patients with type 2 diabetes. Methods: A total of 7 174 type 2 diabetic patients included in National Basic Public Health Service Program in Changshu of Jiangsu Province were recruited as participants. Long-term glucose variability was assessed using standard deviation (SD), coefficient of variation (CV), average real variability (ARV), and variability independent of the mean (VIM) across FPG measurements at the more than three visits. Death information were mainly obtained from the death registry system in Jiangsu. Then Cox proportional hazards regression models were used to estimate the associations of four variability indicators and all-cause mortality's hazard ratios (HRs) and their 95%CIs. Results: Among 55 058.50 person-years of the follow-up, the mean follow-up time was 7.67 years, and 898 deaths occurred during the follow-up period. After adjustment, compared with T1 group, the Cox regression model showed that HRs of T3 group in SD, CV, ARV and VIM were 1.24 (95%CI: 1.03-1.49), 1.20 (95%CI: 1.01-1.43), 1.28 (95%CI: 1.07-1.55) and 1.20 (95%CI:1.01-1.41), respectively. HRs of per 1 SD higher SD, CV, ARV and VIM were 1.13 (95%CI: 1.06-1.21), 1.08 (95%CI: 1.01-1.15), 1.05 (95%CI: 1.00-1.12) and 1.09 (95%CI: 1.02-1.16) for all-cause mortality, respectively. In the stratified analysis, age, gender, hypoglycemic agent and insulin uses had no effect on the above associations (all P for interaction >0.05). Conclusion: Long-term FPG glycemic variability was positively associated with the risk of all-cause mortality in type 2 diabetes patients.

[Characteristics of Guyton's exaggerated forced duction test (FDT) and torsional FDT in congenital superior oblique palsy].

Objective: To investigate the characteristics of Guyton's exaggerated forced duction test (FDT) and torsional FDT in patients with congenital superior oblique palsy (CSOP) and their correlation with clinical features. Methods: This cross-sectional study included single-eye CSOP patients and intermittent exotropia (IXT) patients scheduled for strabismus correction surgery at Tianjin Eye Hospital from September 2021 to March 2022. Prior to surgery, measurements of fovea-disc angle (FDA) and maximum cross-sectional area of the superior oblique muscle (max-CSA) were obtained in both eyes of the patients. The Guyton's exaggerated FDT and torsional FDT were performed intraoperatively to assess the degree of superior oblique muscle relaxation. The characteristics of the two FDT tests and their correlation with vertical strabismus angle, FDA, and max-CSA were analyzed. Statistical analyses were conducted using t-test, ANOVA, Tukey's test, Mann-Whitney U test, and chi-square test. Results: A total of 42 patients (84 eyes) were included in the study, including 19 IXT patients (38 eyes) and 23 CSOP patients (46 eyes, 23 eyes with palsy and 23 eyes without palsy). There were no statistically significant differences in gender composition or age between the IXT and CSOP patients (all P>0.05). The degrees of superior oblique muscle relaxation measured by the Guyton's exaggerated FDT were (-2.52±1.20), (-0.35±0.71), and (-0.03±0.16) for the palsy eye, non-palsy eye, and IXT eyes, respectively, showing significant differences (F=88.10, P<0.001). The torsional FDT measurements yielded external rotation angles of 48.70°±9.67°, 37.39°±5.40°, and 38.95°±2.88° for the palsy eye, non-palsy eye, and IXT eyes, respectively, showing significant differences (F=16.67, P<0.001). There was no statistically significant difference in internal rotation angles (F=2.36, P=0.100). The FDA values were-12.11°±7.42° for IXT patients and-19.02°±4.95° for CSOP patients, while the max-CSA values for the palsy eye and non-palsy eye of CSOP patients were (7.59±4.69) mm² and (11.63±3.64) mm², respectively, all showing significant differences (all P<0.001). The degree of superior oblique muscle tendon relaxation assessed by the Guyton's exaggerated FDT was negatively correlated with the external rotation angle measured by the torsional FDT (r=-0.64, P=0.001). They were positively correlated with max-CSA (r=0.45, P=0.030) and negatively correlated with max-CSA (r=-0.52, P=0.011). However, there was no correlation with vertical and rotational strabismus angle (r=-0.12, P=0.579; r=0.33, P=0.126) and FDA (r=-0.02, P=0.921; r=-0.23, P=0.309). Conclusions: Guyton's exaggerated FDT and torsional FDT can both assess the degree of superior oblique muscle relaxation in patients with CSOP. Furthermore, these two tests are correlated with changes in superior oblique muscle morphology. However, FDT cannot reflect the degree of vertical and rotational strabismus in patients.

Partial cellular reprogramming stably restores the stemness of senescent epidermal stem cells.

OBJECTIVE: Adult stem cell senescence and exhaustion are important drivers of organismal age. Restored stem cell self-renewal has revealed novel therapeutic targets for decreasing the incidence of age-associated diseases (AADs) and prolonging the human health span. Transient ectopic expression of the reprogramming factors Oct3/4, Sox2, Klf4 and c-Myc (collectively known as OSKM) in somatic cells can induce partial cellular reprogramming and effectively ameliorate their age-associated hallmarks. However, how this form of rejuvenation is applied to senescent stem cells remains unknown. MATERIALS AND METHODS: The Integrin-α6highCD71high epidermal stem cells (ESCs) with low self-renewal ability were sorted by flow cytometry and then treated by the interrupted reprogramming induced by transient expression of OSKM. The ability of secondary clones' generation and self-proliferation in vitro, as well as stem cell marker p63, were detected to determine their self-renewal ability. Besides, gene and protein of epidermal cell markers were detected to determine whether their cell identities were retained. Finally, DNA methylation age (eAge) and DNA dehydroxymethylase/methyltransferase were analyzed to explore the alternation of their global DNA methylation pattern during this rejuvenation. RESULTS: The partial reprogramming restored the youthful self-renewal and proliferation in senescent ESCs, including larger secondary clone generation, higher expression of stem cell marker p63 and proliferation marker Ki67, and faster proliferation speed, in each case without abolishing epithelial cellular identity. Moreover, the rejuvenation of adult stem cells could be maintained for 2 weeks after reprogramming factor withdrawal, which was more stable than that of differentiated somatic cells. Additionally, we found that partial reprogramming counteracted the acceleration of eAge in senescent epidermal stem cells and DNA methyltransferase 1 (DNMT1) may play a crucial role in this process. CONCLUSIONS: Partial reprogramming has high therapeutic potential for reversing adult stem cell age, providing an advanced way to treat AADs.

[The distribution of blood pressure and associated factors of the elderly with type 2 diabetes in Jiangsu Province].

Objective: To investigate the distribution of blood pressure and analyze the associated factors of blood pressure of the elderly with type 2 diabetes in Jiangsu Province. Methods: The elderly over 60 years old participants with type 2 diabetes in the communities of Huai'an City and Changshu City, Jiangsu Province were selected in this study. They were divided into two groups: taking antihypertensive drugs and not taking antihypertensive drugs. The demographic characteristics, such as age and sex, and relevant factors were collected by questionnaire. The systolic blood pressure (SBP) and diastolic blood pressure (DBP) were measured by physical examination. The percentile of SBP and DBP in each age group of men and women were described. The kernel density estimation curve was used to show the blood pressure distribution. The trend of blood pressure with age was fitted by locally weighted regression. The logistic regression model was used to analyze relevant factors of blood pressure. Results: A total of 12 949 participants were included in this study, including 7 775 patients in the antihypertensive drug group and 5 174 patients in the group without antihypertensive drugs. The SBP of participants was concentrated at 140-160 mmHg, and their DBP was concentrated at 75-85 mmHg. There were significant differences in the distribution of blood pressure among the subgroups of body mass index (BMI) and rural areas whether taking antihypertensive drugs and not. For participants aged under 80 years old, the SBP showed an increasing trend with age and the DBP showed a decreasing trend with age. Age, BMI ≥24 kg/m2, fasting blood glucose ≥7.0 mmol/L, living in rural areas and no smoking were influencing factors of the elevated SBP; BMI ≥24 kg/m2, male, living in rural areas, no smoking, drinking alcohol and not receiving drug hypoglycemic treatment were influencing factors of the elevated DBP. Conclusion: The SBP of older diabetic adults in Jiangsu Province is at a high level, and the distribution of blood pressure is significantly different between men and women in taking antihypertensive drugs group. The SBP presents a rising trend and the DBP is decreasing at the age of 60-80 years. The blood pressure level of this population are mainly affected by age, BMI, urban and rural areas, smoking.

[Rescue balloon pulmonary angioplasty for a patient with acute exacerbation of chronic thromboembolic pulmonary hypertension supported by extracorporeal membrane oxygenation: a case report].

A 33-year-old male patient was admitted to hospital because of "dyspnea after activity for 3 years and aggravation for 15 days". With a history of membranous nephropathy, irregular anticoagulation led to acute exacerbation of Chronic thromboembolic pulmonary hypertension(CTEPH) and acute respiratory failure, and endotracheal intubation and mechanical ventilation was given. Although treated with thrombolysis and adequate anticoagulation, the condition worsened and hemodynamics deteriorated, and then VA-ECMO was performed. Due to severe pulmonary hypertension and right heart failure,ECMO could not be weaned off, and the patient subsequently developed pulmonary infection, right lung hemorrhage, hyperbilirubinemia, coagulation dysfunction and other complications. Then the patient was transferred to our hospital by airplane, and multidisciplinary discussions were quickly arranged after admission. Considering that the patient was critically ill and complicated with multiple organ failure, pulmonary endarterectomy (PEA) could not be tolerated, rescue balloon pulmonary angioplasty (BPA) was recommended and performed on the second day after admission. The mean pulmonary artery pressure was 59 mmHg(1 mmHg=0.133 kPa) measured by right heart catheterization, and pulmonary angiography showed that the main pulmonary artery was dilated, while the right lower pulmonary artery was completely occluded, and there were multiple stenoses in the branches of the right upper lobe, middle lobe pulmonary artery and the left pulmonary artery. BPA was performed on a total of 9 pulmonary arteries. VA-ECMO was weaned off on day 6 after admission, and the mechanical ventilation was weaned off on day 41 after admission. The patient was successfully discharged on day 72 after admission. Rescue BPA was an effective treatment for severe CTEPH patients who could not be treated with PEA.

[Endoscopic resection of type D trigeminal schwannoma through nasal sinus approach].

Objective: To examine the feasibility and surgical approach of removing type D trigeminal schwannoma through nasal cavity and nasal sinus under endoscope. Methods: Eleven patients with trigeminal schwannoma who were treated in the Department of Otorhinolaryngology, Qilu Hospital of Shandong University from December 2014 to August 2021 were analyzed retrospectively in this study. There were 7 males and 4 females, aged (47.5±13.5) years (range: 12 to 64 years). The neoplasm involved the pterygopalatine fossa, infratemporal fossa, ethmoidal sinus, sphenoid sinus, cavernous sinus, and middle cranial fossa. The size of tumors were between 1.6 cm×2.0 cm×2.0 cm and 5.7 cm×6.0 cm×6.0 cm. Under general anesthesia, the tumors were resected through the transpterygoid approach in 4 cases, through the prelacrimal recess approach in 4 cases, through the extended prelacrimal recess approach in 2 cases, and through the endoscopic medial maxillectomy approach in 1 case. The nasal endoscopy and imaging examination were conducted to detect whether neoplasm recurred or not, and the main clinical symptoms during follow-up. Results: All the surgical procedures were performed under endonasal endoscope, including Gross total resection in 10 patients. The tumor of a 12-year-old patient was not resected completely due to huge tumor size and limited operation space. One patient was accompanied by two other schwannomas located in the occipital region and the ipsilateral parotid gland region originating from the zygomatic branch of the facial nerve, both of which were removed concurrently. After tumor resection, the dura mater of middle cranial fossa was directly exposed in the nasal sinus in 2 cases, including 1 case accompanied by cerebrospinal fluid leakage which was reconstructed by a free mucosal flap obtained from the middle turbinate, the other case was packed by the autologous fat to protect the dura mater. The operation time was (M(IQR)) 180 (160) minutes (range: 120 to 485 minutes). No complications and deaths were observed. No recurrence was observed in the 10 patients with total tumor resection during a 58 (68) months' (range: 10 to 90 months) follow-up. No obvious change was observed in the facial appearance of all patients during the follow-up. Conclusion: Type D trigeminal schwannoma involving pterygopalatine fossa and infratemporal fossa can be removed safely through purely endoscopic endonasal approach by selecting the appropriate approach according to the size and involvement of the tumor.

[Analysis of 42 cases of childhood superior vena cava syndrome associated with mediastinal malignancy].

Objective: To summarize the clinical features, management and outcome of superior vena cava syndrome (SVCS) associated with mediastinal malignancy in children. Methods: Clinical data of 42 children of SVSC associated with mediastinal malignancy in Shanghai Children's Medical Center from January 2015 to December 2021 were collected and analyzed retrospectively. The clinical manifestations, pathological diagnosis, disease diagnosis process, and prognosis were summarized. Results: Among 42 children of SVCS associated with mediastinal malignancy, there were 31 males and 11 females. The age at diagnosis was 8.5 (1.9, 14.9) years. Cough and wheezing (33 cases, 79%), orthopnea (19 cases, 45%) and facial edema (18 cases, 43%) occurred most commonly. T-cell lymphoblastic lymphoma (T-LBL) was the most frequent pathological diagnosis (25 cases, 60%), followed by T-cell acute lymphoblastic leukemia (T-ALL) (7 cases, 17%), anaplastic large cell lymphoma (4 cases, 10%) and diffuse large B-cell lymphoma (2 cases, 5%), peripheral T-lymphoma, Hodgkin lymphoma, Ewing's sarcoma and germ cell tumor (1 case each). Pathological diagnosis was confirmed by bone marrow aspiration or thoracentesis in 14 cases, peripheral lymph node biopsy in 6 cases, and mediastinal biopsy in 22 cases. Twenty-seven cases (64%) had local anesthesia. Respiratory complications due to mediastinal mass developed in 3 of 15 cases who received general anesthesia. Of the 42 cases, 27 cases had sustained remission, 1 case survived with second-line therapy after recurrence, and 14 cases died (2 cases died of perioperative complications and 12 cases died of recurrence or progression of primary disease). The follow-up time was 36.7 (1.2, 76.1) months for 27 cases in continuous complete remission. The 3-year overall survival (OS) and events free survival (EFS) rates of 42 children were 59% (95%CI 44%-79%) and 58% (95%CI 44%-77%) respectively. Conclusions: SVCS associated with mediastinal malignancy in children is a life-threatening tumor emergency with high mortality. The most common primary disease is T-LBL. The most common clinical symptoms and signs are cough, wheezing, orthopnea and facial edema. Clinical management should be based on the premise of stable critical condition and confirm the pathological diagnosis through minimal invasive operation.

[Progress in clinical diagnosis and treatment of multiple primary lung cancer].

With the application of high-resolution chest imaging system and lung cancer screening program, patients with multiple primary lung cancer (MPLC) are becoming a growing population in clinical practice. However, the diagnostic criteria of MPLC and its differentiation from intrapulmonary metastasis of lung cancer (IM) are still controversial, especially in cases with similar histology. On the basis of reviewing the existing literature, this paper discusses the changes of the diagnostic criteria of MPLC and the differential diagnosis methods of imaging, histology and molecular genetics of MPLC and IM, and briefly introduces the application of multidisciplinary diagnosis, algorithm, predictive model and artificial intelligence in the differential diagnosis of MPLC. In addition, we also discuss the latest progress in the treatment of MPLC. Radical surgery is the main method for the treatment of MPLC. Stereotactic body radiation therapy (SBRT) is safe and feasible for inoperable MPLC patients, and targeted therapy and immunotherapy can also be used in MPLC after appropriate patient selection.

[Effects of transient receptor potential vanilloid type 4-specific activator on human vascular endothelial cell functions and blood supply of rat perforator flap and its mechanism].

Objective: To analyze the effects of transient receptor potential vanilloid type 4 (TRPV4) activation on the function and endothelial-to-mesenchymal transition (EndMT) of human umbilical vein endothelial cells (HUVECs), as well as to explore the effects of TRPV4 activation on blood perfusion and survival of rat perforator flap and the mechanism. Methods: The experimental research methods were used. The 3rd to 6th passages of HUVECs were used for experiments and divided into 0.5 µmol/L 4α-phorbol 12, 13-didecanoate (4αPDD) group, 1.0 µmol/L 4αPDD group, 3.0 µmol/L 4αPDD group, 10.0 µmol/L 4αPDD group, and phosphate buffer solution (PBS) group, which were cultivated in corresponding final molarity of 4αPDD and PBS, respectively. The cell proliferation activity at 6 and 12 h of culture was detected using cell counting kit-8 (CCK-8). Another batch of cells was acquired and divided into PBS group, 1 µmol/L 4αPDD group, and 3 µmol/L 4αPDD group, which were treated similarly as described before and then detected for cell proliferation activity at 6, 12, 24, and 48 h of culture. The residual scratch area of cells at post scratch hour (PSH) 12, 24, and 48 was detected by scratch test, and the percentage of the residual scratch area was calculated. The number of migrated cells at 24 and 48 h of culture was detected by Transwell experiment. The tube-formation assay was used to measure the number of tubular structures at 4 and 8 h of culture. The protein expressions of E-cadherin, N-cadherin, Slug, and Snail at 24 h of culture were detected by Western blotting. All the sample numbers in each group at each time point in vitro experiments were 3. A total of 36 male Sprague-Dawley rats aged 8 to 10 weeks were divided into delayed flap group, 4αPDD group, and normal saline group according to the random number table, with 12 rats in each group, and iliolumbar artery perforator flap models on the back were constructed. The flap surgical delay procedure was only performed in the rats in delayed flap group one week before the flap transfer surgery. Neither rats in 4αPDD group nor normal saline group had flap surgical delay; instead, they were intraperitoneally injected with 4αPDD and an equivalent mass of normal saline, respectively, at 10 min before, 24 h after, and 48 h after the surgery. The general state of flap was observed on post surgery day (PSD) 0 (immediately), 1, 4, and 7. The flap survival rates were assessed on PSD 7. The flap blood perfusion was detected by laser speckle contrast imaging technique on PSD 1, 4, and 7. The microvascular density in the flap's choke vessel zone was detected by immunohistochemical staining. All the sample numbers in each group at each time point in vivo experiments were 12. Data were statistically analyzed with analysis of variance for factorial design, analysis of variance for repeated measurement, one-way analysis of variance, least significant difference t test, and Bonferroni correction. Results: At 6 and 12 h of culture, there were no statistically significant differences in cell proliferation activity in the overall comparison among PBS group, 0.5 µmol/L 4αPDD group, 1.0 µmol/L 4αPDD group, 3.0 µmol/L 4αPDD group, and 10.0 µmol/L 4αPDD group (P>0.05). At 6, 12, 24, and 48 h of culture, there were no statistically significant differences in cell proliferation activity in the overall comparison among PBS group, 1 µmol/L 4αPDD group, and 3 µmol/L 4αPDD group (P>0.05). At PSH 12, the percentages of the residual scratch area of cells in 1 µmol/L 4αPDD group and 3 µmol/L 4αPDD group were close to that in PBS group (P>0.05). At PSH 24 and 48, compared with those in PBS group, the percentages of the residual scratch area of cells in 3 µmol/L 4αPDD group were significantly decreased (with t values of 2.83 and 2.79, respectively, P<0.05), while the percentages of the residual scratch area of cells in 1 µmol/L 4αPDD group showed no significant differences (P>0.05). At 24 h of culture, the number of migrated cells in 1 µmol/L 4αPDD group and 3 µmol/L 4αPDD group were close to that in PBS group (P>0.05). At 48 h of culture, the number of migrated cells in 1 µmol/L 4αPDD group and 3 µmol/L 4αPDD groups were significantly greater than that in PBS group (with t values of 6.20 and 9.59, respectively, P<0.01). At 4 h of culture, the numbers of tubular structures of cells in 1 µmol/L 4αPDD group and 3 µmol/L 4αPDD group were significantly greater than that in PBS group (with t values of 4.68 and 4.95, respectively, P<0.05 or <0.01). At 8 h of culture, the numbers of tubular structures of cells in 1 µmol/L 4αPDD and 3 µmol/L 4αPDD groups were similar to that in PBS group (P>0.05). At 24 h of culture, compared with those in PBS group, the protein expression level of E-cadherin of cells in 3 µmol/L 4αPDD group was significantly decreased (t=5.13, P<0.01), whereas there was no statistically significant difference in the protein expression level of E-cadherin of cells in 1 µmol/L 4αPDD group (P>0.05); the protein expression level of N-cadherin of cells in 3 µmol/L 4αPDD group was significantly increased (t=4.93, P<0.01), whereas there was no statistically significant difference in the protein expression level of N-cadherin of cells in 1 µmol/L 4αPDD group (P>0.05); the protein expression levels of Slug of cells in 1 µmol/L 4αPDD group and 3 µmol/L 4αPDD group were significantly increased (with t values of 3.85 and 6.52, respectively, P<0.05 or P<0.01); and the protein expression level of Snail of cells in 3 µmol/L 4αPDD group was significantly increased (t=4.08, P<0.05), whereas there was no statistically significant difference in the protein expression level of Snail of cells in 1 µmol/L 4αPDD group (P>0.05). There were no statistically significant differences in the protein expression levels of E-cadherin, N-cadherin, Slug, or Snail of cells between 1 µmol/L 4αPDD group and 3 µmol/L 4αPDD group (P>0.05). The general condition of flaps of rats in the three groups was good on PSD 0. On PSD 1, the flaps of rats in the three groups were basically similar, with bruising and swelling at the distal end. On PSD 4, the swelling of flaps of rats in the three groups subsided, and the distal end turned dark brown and necrosis occurred, with the area of necrosis in flaps of rats in normal saline group being larger than the areas in 4αPDD group and delayed flap group. On PSD 7, the necrotic areas of flaps of rats in the 3 groups were fairly stable, with the area of necrosis at the distal end of flap of rats in delayed flap group being the smallest. On PSD 7, the flap survival rates of rats in 4αPDD group ((80±13)%) and delayed flap group ((87±9)%) were similar (P>0.05), and both were significantly higher than (70±11)% in normal saline group (with t values of 2.24 and 3.65, respectively, P<0.05 or P<0.01). On PSD 1, the overall blood perfusion signals of rats in the 3 groups were basically the same, and the blood perfusion signals in the choke vessel zone were relatively strong, with a certain degree of underperfusion at the distal end. On PSD 4, the boundary between the surviving and necrotic areas of flaps of rats in the 3 groups became evident, and the blood perfusion signals in the choke vessel zone were improved, with the normal saline group's distal hypoperfused area of flap being larger than the areas in delayed flap group and 4αPDD group. On PSD 7, the blood perfusion signals of overall flap of rats had generally stabilized in the 3 groups, with the intensity of blood perfusion signal in the choke vessel zone and overall flap of rats in delayed flap group and 4αPDD group being significantly greater than that in normal saline group. On PSD 7, the microvascular density in the choke vessel zone of flap of rats in 4αPDD group and delayed flap group were similar (P>0.05), and both were significantly higher than that in normal saline group (with t values of 4.11 and 5.38, respectively, P<0.01). Conclusions: After activation, TRPV4 may promote the migration and tubular formation of human vascular endothelial cells via the EndMT pathway, leading to the enhanced blood perfusion of perforator flap and microvascular density in the choke vessel zone, and therefore increase the flap survival rate.

[Clinical application of expanded internal mammary artery perforator flap combined with vascular supercharge in reconstruction of faciocervical scar].

Objective: To summarize the clinical experience of expanded internal mammary artery perforator (IMAP) flap combined with vascular supercharge in reconstruction of faciocervical scar. Methods: The retrospective observational study was conducted. From September 2012 to May 2021, 23 patients with postburn or posttraumatic faciocervical scars who met the inclusion criteria were admitted to Shanghai Ninth People's Hospital of Shanghai Jiao Tong University School of Medicine, including 18 males and 5 females, aged from 11 to 58 years, all of whom were reconstructed with expanded IMAP flaps. At the first stage, one or two skin and soft tissue expander (s) with appropriate rated capacity were implanted in the anterior chest area according to the location and size of the scars. The IMAP, thoracic branch of supraclavicular artery, and lateral thoracic artery were preserved during the operation. The skin and soft tissue expanders were inflated with normal saline after the operation. The flaps were transferred during the second stage. The dominant IMAP was determined preoperatively using color Doppler ultrasound (CDU) blood flow detector. The faciocervical scars were removed, forming wounds with areas of 9 cm×7 cm-28 cm×12 cm, and the perforators of superficial temporal artery and vein or facial artery and vein were preserved during the operation. The flaps were designed according to the area and size of the wounds after scar resection with the dominant IMAP as the pedicle. Single-pedicle IMAP flaps were used to repair small and medium-sized wounds. For larger defects, the blood perfusion areas of vessels in the anterior chest were evaluated by indocyanine green angiography (ICGA). In situations where the IMAP was insufficient to nourish the entire flap, double-pedicle flaps were designed by using the thoracic branch of supraclavicular artery or lateral thoracic artery for supercharging. Pedicled or free flap transfer was selected according to the distance between the donor areas and recipient areas. After transplantation of flaps, ICGA was conducted again to evaluate blood perfusion of the flaps. The donor sites of flaps were all closed by suturing directly. Statistics were recorded, including the number, rated capacity, normal saline injection volume, and expansion period of skin and soft tissue expanders, the location of the dominant IMAP, the total number of the flaps used, the number of flaps with different types of vascular pedicles, the flap area, the flap survival after the second stage surgery, the occurrence of common complications in the donor and recipient areas, and the condition of follow-up. Results: Totally 25 skin and soft tissue expanders were used in this group of patients, with rated capacity of 200-500 mL, normal saline injection volume of 855-2 055 mL, and expansion period of 4-16 months. The dominant IMAP was detected in the second intercostal space (20 sides) or the third intercostal space (5 sides) before surgery. A total of 25 expanded flaps were excised, including 2 pedicled IMAP flaps, 11 free IMAP flaps, 4 pedicled thoracic branch of supraclavicular artery+free IMAP flaps, and 8 free IMAP+lateral thoracic artery flaps, with flap areas of 10 cm×8 cm-30 cm×14 cm. After the second stage surgery, tip necrosis of flaps in three patients occurred, which healed after routine dressing changes; one patient developed arterial embolism and local torsion on the vascular pedicle at the anastomosis of IMAP and facial artery, and the blood supply recovered after thrombectomy and vascular re-anastomosis. Fourteen patients underwent flap thinning surgery in 1 month to 6 months after the second stage surgery. The follow-up for 4 months to 9 years showed that all patients had improved appearances of flaps and functions of face and neck and linear scar in the donor sites of flaps, and one female patient had obvious nipple displacement and bilateral breast asymmetry. Conclusions: The expanded IMAP flap is matched in color and texture with that of the face and neck, and its incision causes little damage to the chest donor sites. When combined with vascular supercharge, a double-pedicle flap can be designed flexibly to further enhance the blood supply and expand the flap incision area, which is a good choice for reconstruction of large faciocervical scar.

[Imaging features of posterior polymorphous corneal dystrophy observed by in vivo confocal microscopy].

Objective: To identify and analyze imaging features of posterior polymorphous corneal dystrophy (PPCD) by in vivo confocal microscopy (IVCM). Methods: This retrospective case series enrolled 27 eyes of 18 patients (including 10 males and 8 females) who were diagnosed with PPCD at the Department of Ophthalmology in Peking University Third Hospital between January 2013 and December 2019. The mean age was (23.61±14.81) years. There were 9 monocular and 9 binocular cases. All patients were examined by slit-lamp biomicroscopy and IVCM. The visual acuity, the mean endothelial cell density, and the images of IVCM were analyzed in all cases. Results: The mean best-corrected visual acuity was 0.76±0.33, and the mean endothelial cell density was (1 723.6±698.3) cells/mm2. The IVCM images of type 1 PPCD (vesicular lesions) showed hyperreflective, placoid or homocentric lesions at the level of the Descemet's membrane, hyporeflective, oval or round lesions at the level of the Descemet's membrane, and hyporeflective, crater-like lesions at the level of the endothelial cell layer. The IVCM images of type 2 PPCD (band lesions) displayed hyperreflective, band lesions and a fibrous strand structure at the level of the Descemet's membrane, hyporeflective, vesicular lesions at the level of the Descemet's membrane, and hyporeflective, trough-and ridge-like lesions at the level of the endothelial cell layer. The IVCM images of type 3 PPCD (geographic placoid opacities) showed loss of the hexagonal features of endothelial cells and epithelial-like cell transformation. Conclusions: PPCD primarily affects the endothelium and Descemet's membrane. IVCM could highlight the special characteristics of PPCD including hyperreflective lesions at the level of the Descemet's membrane, hyporeflective lesions at the level of the endothelial cell layer, and epithelial-like cell transformation of endothelial cells. IVCM is an invaluable tool for clinical diagnosis and dynamic monitoring of PPCD.

[Cell heterogeneity of laryngeal carcinoma and evolution trajectory of epithelial cells].

Objective: To analyze the classification and functions of cell subsets in laryngeal carcinoma and metastatic lymph nodes, and to explore the evolution trajectory of epithelial cells to tumor cells. Methods: Single-cell RNA sequencing was performed on 5 cases of laryngeal cancer, matched metastatic lymph nodes and 3 normal tissues. Patients were admitted to Ningbo Medical Center Lihuili Hospital from October 22, 2019 to December 16, all patients were male, aged 53-70 years old. Cell subsets of the above-mentioned tissues were analyzed by the Seurat, and the biological functions of cell subpopulation were investigated by functional enrichment analysis. Malignant epithelial cells were identified using copy number variation (CNV). The evolutionary trajectory of epithelial cells to cancer cells was analyzed by cell trajectory analysis, and cancerous transitional cells were identified. The highly expressed genes in transitional cells were analyzed by the FindAllMarker of the Seurat and verified by immunohistochemistry. Results: A total of 66 969 high-quality cells were obtained in 9 major clusters: epithelial cells, T cells, B cells, fibroblasts, endothelial cells, myeloid cells, mast cells, plasmacytoid dendritic cells and nerve cells. The first 5 cell clusters were divided into 8, 6, 4, 3 and 2 subgroups, respectively. Four epithelial cell subsets (C0, C1, C2 and C5) were derived from tumor tissues and metastatic lymph nodes, and had high levels of CNV and tumor cell content. Cell trajectory analysis showed that the evolution trajectory of epithelial cells was from normal epithelial subpopulation C4 to early cancerous cell population C0, which differentiated into three major malignant cell subsets C1, C3, and C5. Epithelial cell C0 may represent the transitional cell population of carcinogenesis, and were enriched in biological processes such as epithelial-mesenchymal transformation and angiogenesis. C0 highly expressed sulforaphane (SFN) which may be related to the occurrence and development of cancer. Immunohistochemistry confirmed that SFN was highly expressed in tumor tissues and metastatic lymph nodes compared with paracancerous tissues. Conclusion: Single-cell sequencing may be used to elucidate the diversity of cells and functions in laryngeal carcinoma tissues and metastatic lymph nodes, and cell population C0 plays a key role in the evolution of cells.

Protein kinase C signalling involved in prothoracicotropic hormone-stimulated prothoracic glands in the silkworm, Bombyx mori.

In the present study, the participation of protein kinase C (PKC) signalling in prothoracicotropic hormone (PTTH)-stimulated ecdysteroidogenesis in Bombyx prothoracic glands (PGs) is demonstrated and characterized. PTTH stimulated phosphorylation of a 37-kDa protein in Bombyx PGs both in vitro and in vivo, as recognized by a PKC substrate antibody. Treatment with either A23187 or thapsigargin also stimulated this 37-kDa protein phosphorylation. PTTH-stimulated phosphorylation of the 37-kDa protein was markedly attenuated in the absence of Ca2+ . The phospholipase C (PLC) inhibitor, U73122, greatly inhibited PTTH-stimulated phosphorylation of this protein, indicating the involvement of Ca2+ and PLC. A mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase (MEK) inhibitor (U0126), a phosphoinositide 3-kinase (PI3K) inhibitor (LY294002) and a chemical activator of adenosine 5'-monophosphate-activated protein kinase (AMPK) (5-aminoimidazole-4-carboxamide-1-ß-d-ribofuranoside) did not affect PTTH-stimulated phosphorylation of the 37-kDa protein, implying that ERK and PI3K/AMPK are not the upstream signalling pathways for PKC-dependent protein phosphorylation. The mitochondrial oxidative phosphorylation inhibitors (the uncoupler carbonyl cyanide p-trifluoromethoxyphenylhydrazone and diphenylene iodonium) inhibited PTTH-stimulated phosphorylation of the 37-kDa protein, indicating its redox regulation. Treatment with PKC inhibitors (either calphostin C, chelerythrine C or rottlerin) reduced PTTH-stimulated phosphorylation of the 37-kDa protein. PTTH-stimulated ecdysteroidogenesis was also inhibited by treatment with rottlerin, thus further confirming participation of PKC-dependent phosphorylation in PTTH signalling. From these results, we demonstrated that redox-regulated PTTH-stimulated PKC signalling is involved in ecdysteroid secretion in Bombyx PGs.

[Etiology of severe community-acquired pneumonia in immunocompromised patients].

Objective: To analyze the etiology of severe community-acquired pneumonia (SCAP) in immunocompromised patients, and to investigate the relationship between underlying diseases and infectious microorganisms. Methods: A retrospective analysis was performed on SCAP in immunocompromised patients admitted to the Fourth Department of Respiratory and Critical Medicine (MICU) of China-Japan Friendship Hospital from January 1, 2017 to December 31, 2019. A total of 119 SCAP patients were finally enrolled, including 65 males (54.6%), with an average age of (59.3±14.5) years. The average of Sequential Organ Failure Assessment (SOFA) score was 6.7±3.6 and the acute physiology and chronic health evaluation (APACHE) Ⅱ score was 19.4±6.8. Sixty (50%) of these patients were finally improved and discharged. Long-term glucocorticoid treatment was the main risk factor for immunocompromise. The difference of pathogenic microorganisms between patients with and without structural lung diseases, and the influence of different pathogenic microorganisms on hospital mortality were calculated, respectively. P<0.05 was considered to be statistically significant. Results: In this study, 99 (83.2%) patients were identified to have positive etiological results, and the incidence of concurrent infection was 54.5%. The top three pathogens were Pneumocystis Jiroveci (55.6%), Cytomegalovirus (47.5%) and Aspergillus (23.2%). Staphylococcus aureus was the most common bacterium, followed by Pseudomonas aeruginosa and Klebsiella pneumoniae. The risk of Pneumocystis Jiroveci infection was significantly higher in patients without underlying lung diseases as compared to those with underlying lung diseases (64.3% vs. 44.2%, P = 0.046). The in-hospital mortality was not different among patients infected with different pathogens(all P>0.05), but was higher in those with mixed infections(56.7% vs. 33.9%, P=0.013). Conclusions: Pneumocystis Jiroveci and Cytomegalovirus were the most common pathogens in immunocompromised patients with severe community-acquired pneumonia, and the incidence of Pneumocystis Jiroveci was significantly higher in patients without underlying lung diseases.