[Clinical Multi-features Analysis of Cystic Lung Adenocarcinoma 
and Construction of Invasive Risk Prediction Model].

BACKGROUND: Cystic lung cancer, a special type of lung cancer, has been paid more and more attention. The most common pathological type of cystic lung cancer is adenocarcinoma. The invasiveness of cystic lung adenocarcinoma is vital for the selection of clinical treatment and prognosis. The aim of this study is to analyze the multiple clinical features of cystic lung adenocarcinoma, explore the independent risk factors of its invasiveness, and establish a risk prediction model. METHODS: A total of 129 cases of cystic lung adenocarcinoma admitted to the Department of Thoracic Surgery of the First Affiliated Hospital of Nanjing Medical University from January 2021 to July 2022 were retrospectively analyzed and divided into pre-invasive group [atypical adenomatous hyperplasia (AAH), adenocarcinoma in situ (AIS) and minimally invasive adenocarcinoma (MIA)] and invasive group [invasive adenocarcinoma (IAC)] according to pathological findings. There were 47 cases in the pre-invasive group, including 19 males and 28 females, with an average age of (51.23±14.96) years. There were 82 cases in the invasive group, including 60 males and 22 females, with an average age of (61.27±11.74) years. Multiple clinical features of the two groups were collected, including baseline data, imaging data and tumor markers. Univariate analysis, LASSO regression and multivariate Logistic regression analysis were used to screen out the independent risk factors of the invasiveness of cystic lung adenocarcinoma, and the risk prediction model was established. RESULTS: In univariate analysis, age, gender, smoking history, history of emphysema, neuron-specific enolase (NSE), number of cystic airspaces, lesion diameter, cystic cavity diameter, nodule diameter, solid components diameter, cyst wall nodule, smoothness of cyst wall, shape of cystic airspace, lobulation, short burr sign, pleural retraction, vascular penetration and bronchial penetration were statistically different between the pre-invasive group and invasive groups (P<0.05). The above variables were processed by LASSO regression dimensionality reduction and screened as follows: age, gender, smoking history, NSE, number of cystic airspaces, lesion diameter, cystic cavity diameter, cyst wall nodule, smoothness of cyst wall and lobulation. Then the above variables were included in multivariate Logistic regression analysis. Cyst wall nodule (P=0.035) and lobulation (P=0.001) were found to be independent risk factors for the invasiveness of cystic lung adenocarcinoma (P<0.05). The prediction model was established as follows: P=e^x/(1+e^x), x=-7.927+1.476* cyst wall nodule+2.407* lobulation, and area under the curve (AUC) was 0.950. CONCLUSIONS: Cyst wall nodule and lobulation are independent risk factors for the invasiveness of cystic lung adenocarcinoma, which have certain guiding significance for the prediction of the invasiveness of cystic lung adenocarcinoma.

Laser capture microdissection transcriptome (LCM RNA-seq) reveals BcDFR is a key gene in anthocyanin synthesis of non-heading Chinese cabbage.

BACKGROUND: Purple non-heading Chinese cabbage [Brassica campestris (syn. Brassica rapa) ssp. chinensis] has become popular because of its richness in anthocyanin. However, anthocyanin only accumulates in the upper epidermis of leaves. Further studies are needed to investigate the molecular mechanisms underlying the specific accumulation of it. RESULTS: In this study, we used the laser capture frozen section method (LCM) to divide purple (ZBC) and green (LBC) non-heading Chinese cabbage leaves into upper and lower epidermis parts (Pup represents the purple upper epidermis, Plow represents the purple lower epidermis, Gup represents the green upper epidermis, Glow represents the green lower epidermis). Through transcriptome sequencing, we found that the DIHYDROFLAVONOL 4-REDUCTASE-encoding gene BcDFR, is strongly expressed in Pup but hardly in others (Plow, Gup, Glow). Further, a deletion and insertion in the promoter of BcDFR in LBC were found, which may interfere with BcDFR expression. Subsequent analysis of gene structure and conserved structural domains showed that BcDFR is highly conserved in Brassica species. The predicted protein-protein interaction network of BcDFR suggests that it interacts with almost all functional proteins in the anthocyanin biosynthesis pathway. Finally, the results of the tobacco transient expression also demonstrated that BcDFR promotes the synthesis and accumulation of anthocyanin. CONCLUSIONS: BcDFR is specifically highly expressed on the upper epidermis of purple non-heading Chinese cabbage leaves and regulates anthocyanin biosynthesis and accumulation. Our study provides new insights into the functional analysis and transcriptional regulatory network of anthocyanin-related genes in purple non-heading Chinese cabbage.

BcABF1 Plays a Role in the Feedback Regulation of Abscisic Acid Signaling via the Direct Activation of BcPYL4 Expression in Pakchoi.

Abscisic acid-responsive element-binding factor 1 (ABF1), a key transcription factor in the ABA signal transduction process, regulates the expression of downstream ABA-responsive genes and is involved in modulating plant responses to abiotic stress and developmental processes. However, there is currently limited research on the feedback regulation of ABF1 in ABA signaling. This study delves into the function of BcABF1 in Pakchoi. We observed a marked increase in BcABF1 expression in leaves upon ABA induction. The overexpression of BcABF1 not only spurred Arabidopsis growth but also augmented the levels of endogenous IAA. Furthermore, BcABF1 overexpression in Arabidopsis significantly decreased leaf water loss and enhanced the expression of genes associated with drought tolerance in the ABA pathway. Intriguingly, we found that BcABF1 can directly activate BcPYL4 expression, a critical receptor in the ABA pathway. Similar to BcABF1, the overexpression of BcPYL4 in Arabidopsis also reduces leaf water loss and promotes the expression of drought and other ABA-responsive genes. Finally, our findings suggested a novel feedback regulation mechanism within the ABA signaling pathway, wherein BcABF1 positively amplifies the ABA signal by directly binding to and activating the BcPYL4 promoter.

CT imaging features of lung ground-glass nodule patients with upgraded intraoperative frozen pathology.

PURPOSE: Intraoperative frozen section pathology (FS) is widely used to guide surgical strategies while the accuracy is relatively low. Underestimating the pathological condition may result in inadequate surgical margins. This study aims to identify CT imaging features related to upgraded FS and develop a predictive model. METHODS: Collected data from 860 patients who underwent lung surgery from January to December 2019. We analyzed the consistency rate of FS and categorized the patients into three groups: Group 1 (n = 360) had both FS and Formalin-fixed Paraffin-embedded section (FP) as non-invasive adenocarcinoma (IAC); Group 2 (n = 128) had FS as non-IAC but FP as IAC; Group 3 (n = 372) had both FS and FP as IAC. Clinical baseline characteristics were compared and propensity score adjustment was used to mitigate the effects of these characteristics. Univariate analyses identified imaging features with inter-group differences. A multivariate analysis was conducted to screen independent risk factors for FS upgrade, after which a logistic regression prediction model was established and a receiver operating characteristic (ROC) curve was plotted. RESULTS: The consistency rate of FS with FP was 84.19%. 26.67% of the patients with non-IAC FS diagnosis were upgraded to IAC. The predictive model's Area Under Curve (AUC) is 0.785. Consolidation tumor ratio (CTR) ≤ 0.5 and smaller nodule diameter are associated with the underestimation of IAC in FS. CONCLUSION: CT imaging has the capacity to effectively detect patients at risk of upstaging during FS.

Hierarchical classification of early microscopic lung nodule based on cascade network.

Purpose: Early-stage lung cancer is typically characterized clinically by the presence of isolated lung nodules. Thousands of cases are examined each year, and one case usually contains numerous lung CT slices. Detecting and classifying early microscopic lung nodules is demanding due to their diminutive dimensions and restricted characterization capabilities. Therefore, a lung nodule classification model that performs well and is sensitive to microscopic lung nodules is needed to accurately classify lung nodules. Methods: This paper uses the Resnet34 network as a basic classification model. A new cascade lung nodule classification method is proposed to classify lung nodules into 6 classes instead of the traditional 2 or 4 classes. It can effectively classify six different nodule types including ground-glass and solid nodules, benign and malignant nodules, and nodules with predominantly ground-glass or solid components. Results: In this paper, the traditional multi-classification method and the cascade classification method proposed in this paper were tested using real lung nodule data collected in the clinic. The test results demonstrate that the cascade classification method in this study achieves an accuracy of 80.04%, outperforming the conventional multi-classification approach. Conclusions: Different from the existing methods for categorizing the benign and malignant nature of lung nodules, the approach presented in this paper can classify lung nodules into 6 categories more accurately. At the same time, This paper proposes a rapid, precise, and dependable approach for classifying six distinct categories of lung nodules, which increases the accuracy categorization compared with the traditional multivariate categorization method.

Three-dimensional imaging anatomical research and clinical implications of V1 +2 d in the left upper lung.

BACKGROUND: To investigate the distribution pattern of V1+2 d in the left superior pulmonary vein and its clinical significance. METHODS: A retrospective analysis was conducted using three-dimensional computed tomographic bronchography and angiography (3D-CTBA) data from 500 lung cancer patients. Statistical analyses were performed to evaluate the incidence and drainage patterns of the three sub-branches of V1+2 d, namely V1+2 d1, V1+2 d2 and V1+2 d3. Furthermore, clinical data from 10 patients' lesions involving V1+2 d were reviewed to illustrate the impact of adjacency to V1+2 d on the surgical approach. RESULTS: The incidences of V1+2 d1, V1+2 d2 and V1+2 d3 were 100%, 76.4% and 100% respectively. The relative interlobar distribution sizes of B3 a and B1+2 c and the left upper division (LUD) vein type influenced the incidence of V1+2 d2 (p < 0.05; p < 0.001). V1+2 d2 predominantly occurred in B3 a = B1+2 c and B1+2 c > B3 a patterns. V1+2 d2 was entirely absent in the B3 a > B1+2 c pattern. V1+2 d2 exhibited a higher incidence in both the central vein (CV) type and the noncentral vein (NCV) type when compared to the semi-central vein (SCV) type (100% vs. 100% vs. 64.8%). The most prevalent venous drainage pattern was the three sub-branches of V1+2 d constituting a major trunk to drain (41.2%). All 10 cases with lesions involving V1+2 d successfully underwent sublobar resection with no complications, and the surgical margin was ≥2 cm. CONCLUSIONS: The three sub-branches of V1+2 d exhibit a high incidence with diverse distribution patterns, yet a discernible pattern exists. For inter- or multi-intersegmental nodules involving V1+2 d, combined segmentectomy and subsegmentectomy or combined subsegmentectomy can ensure the safe margin.

[Determination of 10 carbamate pesticide residues in liquid milk by ultra performance liquid chromatography-tandem mass spectrometry with pass-through solid-phase extraction purification].

Carbamates are used in broad-spectrum insecticides and herbicides, and have highly efficient, low-residue, and long-lasting characteristics. However, this type of pesticide exerts mutagenic, teratogenic, carcinogenic, and other adverse effects, and its frequent use can exceed the recommended scope and limits. Research on the determination of carbamate pesticides mainly focuses on foods of plant origin and pays less attention to foods of animal origin. The methods for carbamate determination described in the current national standards have complicated operating procedures and low efficiency. Therefore, highly efficient and accurate methods for carbamate detection in milk must be established. In this work, a rapid method based on pass-through solid-phase extraction (SPE) purification coupled with ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was developed for the simultaneous determination of 10 carbamate pesticides in liquid milk. The pretreatment and instrument methods were systematically optimized. The milk sample was extracted with acetonitrile, and then purified using a Captiva EMR-Lipid filtration kit. The purified extract was separated on an ACQUITY UPLC BEH C18 column with mobile phase of methanol and 0.1% formic acid aqueous solution in gradient elution. The flow rate was 0.3 mL/min. Column temperature was 35 ℃. Quantitative analysis was performed using the external standard method with matrix matching curves. The 10 carbamate pesticides showed good linear relationships in the mass concentration range of 2-200 µg/L, with correlation coefficients greater than 0.999. The limits of detection (LODs) and quantification (LOQs) for the 10 carbamate pesticides were 0.045-0.23 and 0.15-0.77 µg/kg, respectively. Recovery tests were conducted using the blank-matrix method at three spiked levels of 15, 50, and 100 µg/kg, and good recoveries for the 10 carbamate pesticides were obtained. In particular, the recoveries for the three spiked levels of 15, 50, and 100 µg/kg were 68.7%-93.3% with relative standard deviations (RSDs) of 1.8%-8.0%. The proposed method is efficient, convenient, accurate, and suitable for the rapid detection of the 10 carbamate pesticides in liquid milk. Compared with the conventional NH2 and ENVITM-18 SPE columns used in the national standard determination method, the proposed method demonstrated better purification effects. The recoveries for aldicarb sulfoxide, aldicarb sulfone, methomyl, and carbaryl after purification using the Captiva EMR-Lipid kit increased from 60% to 80%. Thus, the proposed method is suitable for targets with strong polarity and gives measurement results with good repeatability and accuracy.

Difficulty of Video-Assisted Thoracoscopic Surgery Segmentectomy: Proposal for a New Classification.

BACKGROUND: Segmentectomy has classically been distinguished as "simple" or "complex" based on the number of intersegmental planes (ISPs) dissected. However, with the increasing variety and complexity of segmentectomies, it is clear that a classification based on the number of ISPs alone is inadequate. This study aimed to develop a new classification to predict the surgical difficulty of video-assisted thoracoscopic surgery (VATS) segmentectomy. METHODS: The study retrospectively reviewed 1868 patients who underwent VATS segmentectomy between January 2014 and December 2019. Uni- and multivariate analyses were performed to identify predictors associated with prolonged operative time (>140 min), and a scoring system was constructed to classify the surgical difficulty of VATS segmentectomy. RESULTS: Altogether, 1868 VATS segmentectomies were divided into three groups: group 1 (low difficulty, including segmentectomy with only one intersegmental plane [ISP] dissection), group 2 (intermediate difficulty, including a single segmentectomy with more than one ISP dissection and a single subsegmentectomy), group 3 (high difficulty level, including combined resection with more than one ISP dissection). This classification effectively differentiated the three groups in terms of operative time, estimated blood loss, major complications, and overall complications (all p < 0.001). For receiver operating characteristic analysis, the new classification showed significantly better differentiation performance in terms of operative time (p < 0.001), estimated blood loss (p = 0.004), major complications (p = 0.002), and overall complications (p = 0.012) than the simple/complex classification. CONCLUSIONS: This new three-level classification accurately predicted the surgical difficulty of VATS segmentectomy.

Inhibiting the transcription and replication of Ebola viruses by disrupting the nucleoprotein and VP30 protein interaction with small molecules.

Ebola virus (EBOV) causes hemorrhagic fever in humans with high morbidity and fatality. Although over 45 years have passed since the first EBOV outbreak, small molecule drugs are not yet available. Ebola viral protein VP30 is a unique RNA synthesis cofactor, and the VP30/NP interaction plays a critical role in initiating the transcription and propagation of EBOV. Here, we designed a high-throughput screening technique based on a competitive binding assay to bind VP30 between an NP-derived peptide and a chemical compound. By screening a library of 8004 compounds, we obtained two lead compounds, Embelin and Kobe2602. The binding of these compounds to the VP30-NP interface was validated by dose-dependent competitive binding assay, surface plasmon resonance, and thermal shift assay. Moreover, the compounds were confirmed to inhibit the transcription and replication of the Ebola genome by a minigenome assay. Similar results were obtained for their two respective analogs (8-gingerol and Kobe0065). Interestingly, these two structurally different molecules exhibit synergistic binding to the VP30/NP interface. The antiviral efficacy (EC50) increased from 1 µM by Kobe0065 alone to 351 nM when Kobe0065 and Embelin were combined in a 4:1 ratio. The synergistic anti-EBOV effect provides a strong incentive for further developing these lead compounds in future studies.

TRPV4 channel is involved in HSV-2 infection in human vaginal epithelial cells through triggering Ca2+ oscillation.

Herpes simplex virus (HSV) infection induces a rapid and transient increase in intracellular calcium concentration ([Ca2+]i), which plays a critical role in facilitating viral entry. T-type calcium channel blockers and EGTA, a chelate of extracellular Ca2+, suppress HSV-2 infection. But the cellular mechanisms mediating HSV infection-activated Ca2+ signaling have not been completely defined. In this study we investigated whether the TRPV4 channel was involved in HSV-2 infection in human vaginal epithelial cells. We showed that the TRPV4 channel was expressed in human vaginal epithelial cells (VK2/E6E7). Using distinct pharmacological tools, we demonstrated that activation of the TRPV4 channel induced Ca2+ influx, and the TRPV4 channel worked as a Ca2+-permeable channel in VK2/E6E7 cells. We detected a direct interaction between the TRPV4 channel protein and HSV-2 glycoprotein D in the plasma membrane of VK2/E6E7 cells and the vaginal tissues of HSV-2-infected mice as well as in phallic biopsies from genital herpes patients. Pretreatment with specific TRPV4 channel inhibitors, GSK2193874 (1-4 µM) and HC067047 (100 nM), or gene silence of the TRPV4 channel not only suppressed HSV-2 infectivity but also reduced HSV-2-induced cytokine and chemokine generation in VK2/E6E7 cells by blocking Ca2+ influx through TRPV4 channel. These results reveal that the TRPV4 channel works as a Ca2+-permeable channel to facilitate HSV-2 infection in host epithelial cells and suggest that the design and development of novel TRPV4 channel inhibitors may help to treat HSV-2 infections.

Efficacy of oxidized regenerated cellulose combined with fibrin glue in reducing pulmonary air leakage after segmentectomy in a porcine lung model.

Objectives: Pulmonary air leakage is a common complication following lung resection. We have designed a new method combining oxidized regenerated cellulose and fibrin glue to cover the intersegmental plane in clinical lung segmentectomy to prevent postoperative air leakage. In this study, an excised porcine lung segmentectomy model was created to validate its adhesive strength and effect on reducing air leakage. Methods: In the pre-experiment, six different larger lung segments were separated using electrocautery on the fresh isolated porcine lungs (n = 5 in each group). The air leakage degree and operation time of the lung segments were comprehensively evaluated to select the most suitable target segment for establishing the ex vivo porcine lung segmentectomy models. In the experiment, according to the different materials covered on the intersegmental plane, these models were randomly divided into four groups: group A used fibrin glue and oxidized regenerated cellulose (ORC) mesh (n = 20); group B used fibrin glue and polyglycolic acid (PGA) sheet (n = 20); group C used fibrin glue (n = 20); group D was the blank control group (n = 20). The minimum air leakage pressure (MALP) of the selected target segment in each group was measured using a stepwise increase of airway pressure, and histological assessment was performed on the sealed area samples from the four groups. Results: The operation time of the a segment of the right cranial lobe (R1a) was shorter than that of other segments (p < 0.05), and there was no significant difference in the air leakage pressures between the six isolated segments (p = 0.76); thus, R1a was chosen for segmentectomy. In addition, the MALP was significantly higher in group A (41.8 ± 4.5 cmH2O) than in groups C (28.1 ± 2.3 cmH2O) and D (17.3 ± 1.2 cmH2O) (both p < 0.001). The MALP of group B (69.5 ± 5.2 cmH2O) was significantly higher than that of group A (p < 0.001), whereas that of group C was significantly higher than that of group D (p < 0.001). Histological examination confirmed that the combined use of fibrin glue and ORC or PGA patch adhered more firmly to the intersegmental plane than that of fibrin glue alone, although some gaps could be seen between the fibrin glue and the surface of the lung segments in group C. Conclusion: The application of ORC combined with fibrin glue on the intersegmental plane has a good sealing performance in the ex vivo porcine lung segmentectomy model, suggesting that ORC may be an effective alternative material to replace PGA sheet to combine with fibrin glue for preventing air leakage after segmentectomy.

[Comparison of Two-dimensional and Three-dimensional Features of Chest CT
 in the Diagnosis of Invasion of Pulmonary Ground Glass Nodules].

BACKGROUND: At present, more and more studies predict invasive adenocarcinoma (IAC) through three-dimensional features of pulmonary nodules, but few studies have confirmed that three-dimensional features have more advantages in diagnosing IAC than traditional two-dimensional features of pulmonary nodules. This study analyzed the differences of chest computed tomography (CT) features between IAC and minimally invasive adenocarcinoma (MIA) from three-dimensional and two-dimensional levels, and compared the ability of diagnosing IAC. The non-invasive adenocarcinoma group includes precursor glandular lesions (PGL) and minimally invasive adenocarcinoma (MIA). METHODS: The clinical data of 1,045 patients with ground glass opacity (GGO) from January to December 2019 were collected. Then the correlation between preoperative CT image characteristics and pathological results were analyzed retrospectively. The independent influencing factors for the identification of IAC were screened out according to two-dimensional and three-dimensional classification by multivariate Logistic regression and the cut-off point for the identification of IAC was found out through the receiver operating characteristic (ROC) curve. At last, the ability of diagnosing IAC was evaluated by Yoden index. RESULTS: The diameter of nodule, the diameter of solid component, the diameter of mediastinal window nodule in two-dimensional factors, and the volume of nodule, the volume of solid part and the average CT value in three-dimensional factors were independent risk factors for the diagnosis of IAC. These factors were arranged by Yoden index: solid partial volume (0.601)>nodule volume (0.536)>solid component diameter (0.525)>nodule diameter (0.518)>mediastinal window nodule diameter (0.488)>proportion of solid component volume (0.471)>1-tumor disappearance ratio (TDR) (0.468)>consolidation tumor ratio (CTR) (0.394)>average CT value (0.380). CONCLUSIONS: The CT features of three-dimensional are better than two-dimensional in the diagnosis of IAC, and the size of solid components is better than the overall size of nodules.

PARP1 Might Substitute HSF1 to Reactivate Latent HIV-1 by Binding to Heat Shock Element.

At present, the barrier to HIV-1 functional cure is the persistence of HIV-1 reservoirs. The "shock (reversing latency) and kill (antiretroviral therapy)" strategy sheds light on reducing or eliminating the latent reservoir of HIV-1. However, the current limits of latency-reversing agents (LRAs) are their toxicity or side effects, which limit their practicability pharmacologically and immunologically. Our previous research found that HSF1 is a key transcriptional regulatory factor in the reversion of HIV-1 latency. We then constructed the in vitro HSF1-knockout (HSF1-KO) HIV-1 latency models and found that HSF1 depletion inhibited the reactivation ability of LRAs including salubrinal, carfizomib, bortezomib, PR-957 and resveratrol, respectively. Furthermore, bortezomib/carfizomib treatment induced the increase of heat shock elements (HSEs) activity after HSF1-KO, suggesting that HSEs participated in reversing the latent HIV-1. Subsequent investigation showed that latent HIV-1-reversal by H2O2-induced DNA damage was inhibited by PARP1 inhibitors, while PARP1 was unable to down-regulate HSF1-depleted HSE activity, indicating that PARP1 could serve as a replaceable protein for HSF1 in HIV-1 latent cells. In summary, we succeeded in finding the mechanisms by which HSF1 reactivates the latent HIV-1, which also provides a theoretical basis for the further development of LRAs that specifically target HSF1.

The New Variation in the Promoter Region of FLOWERING LOCUS T Is Involved in Flowering in Brassica rapa.

Flowering time is an important agronomic trait in Brassica rapa and has a wide range of variation. The change from vegetative to reproductive development is a major transition period, especially in flowering vegetable crops. In this study, two non-heading Chinese cabbage varieties with significantly different flowering times, Pak-choi (B. rapa var. communis Tesn et Lee) and Caitai (B. rapa var. tsaitai Hort.), were used to construct segregated F2 populations. The bulk-segregant approach coupled with whole genome re-sequencing was used for QTL sequencing (QTL-seq) analysis to map flowering time traits. The candidate genes controlling flowering time in B. rapa were predicted by homologous gene alignment and function annotation. The major-effect QTL ft7.1 was detected on chromosome A07 of B. rapa, and the FT family gene BrFT was predicted as the candidate gene. Moreover, a new promoter regional difference of 1577 bp was revealed by analyzing the sequence of the BrFT gene. The promoter region activity analysis and divergent gene expression levels indicated that the difference in the promoter region may contribute to different flowering times. These findings provide insights into the mechanisms underlying the flowering time in Brassica and the candidate genes regulating flowering in production.

Genome-wide identification of LEA gene family and cold response mechanism of BcLEA4-7 and BcLEA4-18 in non-heading Chinese cabbage [Brassica campestris (syn. Brassica rapa) ssp. chinensis].

Cold stress is a key factor limiting the yield and quality of non-heading Chinese cabbage. The hydrophilic protective protein LEA plays an important role in plant abiotic stress. In this study, 72 BcLEAs were identified from non-heading Chinese cabbage and divided into 9 subfamilies by phylogenetic analysis. Gene structure analysis showed that BcLEAs were unevenly distributed on 10 chromosomes, with few introns. Through analyzing the expression of these genes under cold stress by RNA-seq and qRT-PCR, two genes (BcLEA4-7 and BcLEA4-18) highly sensitive to cold stress were identified, whose roles in cold tolerance of non-heading Chinese cabbage were demonstrated by virus-induced gene silencing. The BcLEA promoters were analyzed to study the cold response mechanism of BcLEA4-7 and BcLEA4-18, revealing that both BcLEA4-7 and BcLEA4-18 promoters contained two CRT/DRE elements. Subsequently, it was found that the promoters isolated from non-heading Chinese cabbage could be activated at low temperatures. Further analysis showed BcCBF2 in non-heading Chinese cabbage interacted with two CRT/DRE elements in BcLEA4-7 and BcLEA4-18 promoters to stimulate their activity, indicating that BcCBF2 is an upstream regulator. Meanwhile, the CRT/DRE element located in BcLEA4-7 promoter (-219 bp to -171 bp) and BcLEA4-18 promoter (-234 bp to -186 bp) was more likely to be activated by BcCBF2, which may be attributed to its flanking sequence. These data laid a foundation for further understanding the functional role and regulatory mechanism of BcLEAs in cold stress tolerance.

Clinical significance of the diverse interlobar veins hidden in the upper oblique fissure.

Background: The interlobar veins hidden in the upper oblique fissure (UOF) of the right lung are usually mismanaged cursorily according to the target lobe, which results in accidental injury of the interlobar veins and complications. The detailed classification of interlobar veins based on surgical anatomical analysis is of great clinical significance. Methods: Three-dimensional computed tomography bronchography and angiography (3D-CTBA) reconstructed images of 398 patients from January 2019 to June 2020 were retrospectively analyzed. The interlobar veins in the UOF were observed and classified according to their morphology and distribution. The classification model was further validated in 153 patients who underwent surgery involving dissection of the UOF, and related surgical results were analyzed. Results: The distribution of interlobar veins was diverse, and the general morphology could be divided into 2 main categories and 30 subtypes in the 3D-CTBA images of the 398 patients. Analysis of the 153 patients' surgical data showed that 60 patients suffered from interlobar vein injury. Interlobar veins hidden in an incomplete UOF were the most susceptible to accidental damage (χ2=12.856, P=0.020). A receiver operating characteristic (ROC) curve analysis showed that an interlobar vein diameter larger than 2.4 mm for the oblique fissure interlobar vein type or less than 2 mm for the mediastinal interlobar vein type was associated with a higher risk of injuries (P<0.001). Conclusions: The diversity of interlobar veins and the completeness of the UOF were noteworthy risk factors in surgery involving dissection of the UOF.

[Relationship between EGFR, ALK Gene Mutation and Imaging 
and Pathological Features in Invasive Lung Adenocarcinoma].

BACKGROUND: At present, the research progress of targeted therapy for epidermal growth factor receptor (EGFR) and anaplastic lymphoma kinase (ALK) gene mutations in lung adenocarcinoma is very rapid, which brings new hope for the treatment of advanced lung adenocarcinoma patients. However, the specific imaging and pathological features of EGFR and ALK gene mutations in adenocarcinoma are still controversial. This study will further explore the correlation between EGFR, ALK gene mutations and imaging and pathological features in invasive lung adenocarcinoma. METHODS: A total of 525 patients with lung adenocarcinoma who underwent surgery in our center from January 2018 to December 2019 were included. According to the results of postoperative gene detection, the patients were divided into EGFR gene mutation group, ALK gene mutation group and wild group, and the EGFR gene mutation group was divided into exon 19 and exon 21 subtypes. The pathological features of the mutation group and wild group, such as histological subtype, lymph node metastasis, visceral pleural invasion (VPI) and imaging features such as tumor diameter, consolidation tumor ratio (CTR), lobulation sign, spiculation sign, pleural retraction sign, air bronchus sign and vacuole sign were analyzed by univariate analysis and multivariate Logistic regression analysis to explore whether the gene mutation group had specific manifestations. RESULTS: EGFR gene mutation group was common in women (OR=2.041, P=0.001), with more pleural traction sign (OR=1.506, P=0.042), and had little correlation with lymph node metastasis and VPI (P>0.05). Among them, exon 21 subtype was more common in older (OR=1.022, P=0.036), women (OR=2.010, P=0.007), and was associated with larger tumor diameter (OR=1.360, P=0.039) and pleural traction sign (OR=1.754, P=0.029). Exon 19 subtype was common in women (OR=2.230, P=0.009), with a high proportion of solid components (OR=1.589, P=0.047) and more lobulation sign (OR=2.762, P=0.026). ALK gene mutations were likely to occur in younger patients (OR=2.950, P=0.045), with somking history (OR=1.070, P=0.002), and there were more micropapillary components (OR=4.184, P=0.019) and VPI (OR=2.986, P=0.034) in pathology. CONCLUSIONS: The EGFR and ALK genes mutated adenocarcinomas have specific imaging and clinicopathological features, and the mutations in exon 19 or exon 21 subtype have different imaging features, which is of great significance in guiding the clinical diagnosis and treatment of pulmonary nodules.

A Five-Helix-Based SARS-CoV-2 Fusion Inhibitor Targeting Heptad Repeat 2 Domain against SARS-CoV-2 and Its Variants of Concern.

The prolonged duration of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pandemic has resulted in the continuous emergence of variants of concern (VOC, e.g., Omicron) and variants of interest (VOI, e.g., Lambda). These variants have challenged the protective efficacy of current COVID-19 vaccines, thus calling for the development of novel therapeutics against SARS-CoV-2 and its VOCs. Here, we constructed a novel fusion inhibitor-based recombinant protein, denoted as 5-Helix, consisting of three heptad repeat 1 (HR1) and two heptad repeat 2 (HR2) fragments. The 5-Helix interacted with the HR2 domain of the viral S2 subunit, the most conserved region in spike (S) protein, to block homologous six-helix bundle (6-HB) formation between viral HR1 and HR2 domains and, hence, viral S-mediated cell-cell fusion. The 5-Helix potently inhibited infection by pseudotyped SARS-CoV-2 and its VOCs, including Delta and Omicron variants. The 5-Helix also inhibited infection by authentic SARS-CoV-2 wild-type (nCoV-SH01) strain and its Delta variant. Collectively, our findings suggest that 5-Helix can be further developed as either a therapeutic or prophylactic to treat and prevent infection by SARS-CoV-2 and its variants.