Investigating the cis-regulatory basis of C3 and C4 photosynthesis in grasses at single-cell resolution.

While considerable knowledge exists about the enzymes pivotal for C4 photosynthesis, much less is known about the cis-regulation important for specifying their expression in distinct cell types. Here, we use single-cell-indexed ATAC-seq to identify cell-type-specific accessible chromatin regions (ACRs) associated with C4 enzymes for five different grass species. This study spans four C4 species, covering three distinct photosynthetic subtypes: Zea mays and Sorghum bicolor (NADP-dependent malic enzyme), Panicum miliaceum (NAD-dependent malic enzyme), Urochloa fusca (phosphoenolpyruvate carboxykinase), along with the C3 outgroup Oryza sativa. We studied the cis-regulatory landscape of enzymes essential across all C4 species and those unique to C4 subtypes, measuring cell-type-specific biases for C4 enzymes using chromatin accessibility data. Integrating these data with phylogenetics revealed diverse co-option of gene family members between species, showcasing the various paths of C4 evolution. Besides promoter proximal ACRs, we found that, on average, C4 genes have two to three distal cell-type-specific ACRs, highlighting the complexity and divergent nature of C4 evolution. Examining the evolutionary history of these cell-type-specific ACRs revealed a spectrum of conserved and novel ACRs, even among closely related species, indicating ongoing evolution of cis-regulation at these C4 loci. This study illuminates the dynamic and complex nature of cis-regulatory elements evolution in C4 photosynthesis, particularly highlighting the intricate cis-regulatory evolution of key loci. Our findings offer a valuable resource for future investigations, potentially aiding in the optimization of C3 crop performance under changing climatic conditions.

Chromatin and regulatory differentiation between bundle sheath and mesophyll cells in maize.

C4 plants partition photosynthesis enzymes between the bundle sheath (BS) and the mesophyll (M) cells for the better delivery of CO2 to RuBisCO and to reduce photorespiration. To better understand how C4 photosynthesis is regulated at the transcriptional level, we performed RNA-seq, ATAC-seq, ChIP-seq and Bisulfite-seq (BS-seq) on BS and M cells isolated from maize leaves. By integrating differentially expressed genes with chromatin features, we found that chromatin accessibility coordinates with epigenetic features, especially H3K27me3 modification and CHH methylation, to regulate cell type-preferentially enriched gene expression. Not only the chromatin-accessible regions (ACRs) proximal to the genes (pACRs) but also the distal ACRs (dACRs) are determinants of cell type-preferentially enriched expression. We further identified cell type-preferentially enriched motifs, e.g. AAAG for BS cells and TGACC/T for M cells, and determined their corresponding transcription factors: DOFs and WRKYs. The complex interaction between cis and trans factors in the preferential expression of C4 genes was also observed. Interestingly, cell type-preferentially enriched gene expression can be fine-tuned by the coordination of multiple chromatin features. Such coordination may be critical in ensuring the cell type-specific function of key C4 genes. Based on the observed cell type-preferentially enriched expression pattern and coordinated chromatin features, we predicted a set of functionally unknown genes, e.g. Zm00001d042050 and Zm00001d040659, to be potential key C4 genes. Our findings provide deep insight into the architectures associated with C4 gene expression and could serve as a valuable resource to further identify the regulatory mechanisms present in C4 species.

Characterization of regulatory modules controlling leaf angle in maize.

Leaf angle is an important agronomic trait determining maize (Zea mays) planting density and light penetration into the canopy and contributes to the yield gain in modern maize hybrids. However, little is known about the molecular mechanisms underlying leaf angle beyond the ZmLG1 (liguleless1) and ZmLG2 (Liguleless2) genes. In this study, we found that the transcription factor (TF) ZmBEH1 (BZR1/BES1 homolog gene 1) is targeted by ZmLG2 and regulates leaf angle formation by influencing sclerenchyma cell layers on the adaxial side. ZmBEH1 interacted with the TF ZmBZR1 (Brassinazole Resistant 1), whose gene expression was also directly activated by ZmLG2. Both ZmBEH1 and ZmBZR1 are bound to the promoter of ZmSCL28 (SCARECROW-LIKE 28), a third TF that influences leaf angle. Our study demonstrates regulatory modules controlling leaf angle and provides gene editing targets for creating optimal maize architecture suitable for dense planting.

Integrative genomic and transcriptomic analysis reveals immune subtypes and prognostic markers in ovarian clear cell carcinoma.

BACKGROUND: We performed an integrative genomic and transcriptomic profiling to identify molecular subtypes and prognostic markers with special focus on immune-related pathways. METHODS: Totally, 50 Chinese patients were subjected to targeted next-generation sequencing and transcriptomic sequencing. RESULTS: Two distinct subgroups were identified as immune (22.0%) and non-immune (78.0%) based on the immune-pathway related hierarchical clustering. Surprisingly, patients with immune subtype had a significantly worse survival. The prognostic capacity was validated in external cohorts. The immune group had higher expression of genes involved in pro-inflammation and checkpoints. PD-1 signalling pathway was enriched in the immune subtype. Besides, the immune cluster presented enriched expression of genes involved in epithelial-mesenchymal transition, angiogenesis and PI3K-AKT-mTOR signalling, while the non-immune subtype had higher expression of metabolic pathways. The immune subtype had a higher mutation rate of PIK3CA though significance was not achieved. Lastly, we established a prognostic immune signature for overall survival. Interestingly, the immune signature could also be applied to renal clear cell carcinoma, but not to other histologic subtype of ovarian cancer. CONCLUSIONS: An immune subtype of OCCC was identified with poor survival and enrichment of PD-1 and PI3K-AKT-mTOR signalling. We constructed and validated a robust prognostic immune signature of OCCC patients.

Feedback regulation of auxin signaling through the transcription of H2A.Z and deposition of H2A.Z to SMALL AUXIN UP RNAs in Arabidopsis.

Auxin is a critical phytohormone that is involved in the regulation of most plant growth and developmental responses. In particular, epigenetic mechanisms, like histone modifications and DNA methylation, were reported to affect auxin biosynthesis and transport. However, the involvement of other epigenetic factors, such as histone variant H2A.Z, in the auxin-related developmental regulation remains unclear. We report that the histone variant H2A.Z knockdown mutant in Arabidopsis Col-0 ecotype, h2a.z-kd, has more lateral roots and weak gravitational responses related to auxin-regulated growth performances. Further study revealed that auxin promotes the eviction of H2A.Z from the auxin-responsive genes SMALL AUXIN-UP RNAs (SAURs) to activate their transcriptions. We found that IAA promotes the transcription of H2A.Z genes through HOMEOBOX PROTEIN 22/25 (AtHB22/25) transcription factors which work as downstream targets of ARF7/19 in auxin signaling. Double mutant of hb22 hb25 showed similar lateral root and gravitropism phenotypes to h2a.z-kd. Our results shed light on a reciprocal regulation hub through INOSITOL AUXOTROPHY 80-mediated H2A.Z eviction and ARF7/19-HB22/25-mediated H2A.Z transcription to modulate the activation of SAURs and plant growth in Arabidopsis.

Invasive apocrine carcinoma of the breast: clinicopathologic features and comprehensive genomic profiling of 18 pure triple-negative apocrine carcinomas.

Pure invasive apocrine carcinoma is a rare type of primary breast cancer, constituting ~1% of all breast cancers. Since most pure invasive apocrine carcinomas are triple negative, the lack of targeted therapies for triple-negative breast cancer has fostered efforts to discover actionable molecular targets in these tumors. In this study, we analyzed the clinicopathologic characteristics and comprehensive genomic profiling of 18 patients with pure triple-negative apocrine carcinomas (TNACs) using a 324-gene panel assay (FoundationOne CDx). The median age of these patients was 55.5 years, and the postmenopausal status rate was 77.8%. In total, 83.3% of patients were diagnosed with histological grade II, and 16.7% were diagnosed with grade III. The majority of patients presented at an early tumor-node-metastasis (TNM) stage (I: 38.9%; II: 50.0%; and III: 11.1%). The mean Ki-67 index was 9.7%, and the percent of PD-L1 positivity was 11.7%. With a median follow-up period of 76.5 months, one patient died, and two experienced distant metastases. There were 61 clinically relevant genomic alterations among all 18 pure TNACs, and the mean tumor mutation burden (TMB) was 3 Mut/Mb. The top ranked altered genes were PIK3CA (72.2%), PTEN (33.3%) and TP53 (27.8%). There were four novel mutations found in PTEN and an actionable rearrangement involving FGFR2-TACC2 that has not been reported in breast cancer before. In total, 88.9%, 50%, 44.4%, and 16.7% of TNACs had at least one clinically relevant genomic alteration in genes involved in the PI3K/mTOR, cell cycle, RAS/RAF/MEK and growth factor receptor-related pathways, respectively. All patients had at least one clinically relevant genomic alteration, and 94.4% had at least one actionable alteration. To the best of our knowledge, this study is the largest genomic sequencing cohort of pure TNACs. Incorporation of comprehensive genomic profiling into TNACs might shed light on potential therapeutic opportunities for both targeted drugs and immune checkpoint inhibitors.

Intense basolateral membrane staining indicates HER2 positivity in invasive micropapillary breast carcinoma.

Invasive micropapillary carcinoma is characterized by the inside-out growth of tumor clusters and displays incomplete membrane immunostaining of HER2. According to the 2018 American Society of Clinical Oncology and the College of American Pathologists (ASCO/CAP) HER2-testing recommendation, moderate to intense but incomplete staining could be scored as immunohistochemical 2+. Furthermore, the criteria of immunohistochemical 3+ for this staining pattern are not mentioned. One hundred and forty-seven cases of invasive micropapillary carcinoma with moderate-to-intense HER2 immunostaining were enrolled. Invasive micropapillary carcinoma components of all cases were scored as immunohistochemical 2+ based on the 2018 ASCO/CAP recommendation. The invasive micropapillary carcinoma component varied from 10% to 100% (mean, 80%). Invasive micropapillary carcinoma components of all 147 tumors exhibited reversed polarity and incomplete basolateral HER2 membrane staining. One hundred and seventeen of the tumors (80%, 117/147) had moderate staining, and 38 (32%, 38/117) showed HER2 gene amplification by fluorescence in-situ hybridization. HER2 gene was amplified in all the remaining 30 tumors (20%, 30/147) that exhibited intense basolateral membrane staining. Besides, average HER2 signals per cell and ratio of HER2/CEP17 were significantly higher in the intense-staining tumors compared with the moderate-staining tumors (p < 0.0001). Follow-up data were available for 140 patients. None of the patients were died. The follow-up time ranged from 1 month to 99 months (median, 57 months). Thirteen (9%, 13/140) patients exhibited disease progression (recurrence or metastasis). HER2 gene amplification was correlated inversely with estrogen receptor (p = 0.000) and progesterone receptor (p = 0.000) expression, and positively with histological grade (p = 0.003) and disease progression (p = 0.000). Invasive micropapillary carcinoma with intense clear linear basolateral membrane immunostaining indicates HER2 positivity, even if the staining is incomplete. They should be classified as immunohistochemical 3+ rather than immunohistochemical 2+, which would avoid further fluorescence in-situ hybridization-testing procedure and greatly save the related time, labor, and financial costs. Ultimately, ensure all patients with HER2 gene amplification obtain effective targeted therapy in time.

Plant and animal chromatin three-dimensional organization: similar structures but different functions.

Chromatin is the main carrier of genetic information and is non-randomly distributed within the nucleus. Next-generation sequence-based chromatin conformation capture technologies have enabled us to directly examine its three-dimensional organization at an unprecedented scale and resolution. In the best-studied mammalian models, chromatin folding can be broken down into three hierarchical levels, compartment, domains, and loops, which play important roles in transcriptional regulation. Although similar structures have now been identified in plants, they might not possess exactly the same functions as the mammalian ones. Here, we review recent Hi-C studies in plants, compare plant chromatin structures with their mammalian counterparts, and discuss the differences between plants with different genome sizes.

Tissue-specific Hi-C analyses of rice, foxtail millet and maize suggest non-canonical function of plant chromatin domains.

Chromatins are not randomly packaged in the nucleus and their organization plays important roles in transcription regulation, which is best studied in the mammalian models. Using in situ Hi-C, we have compared the 3D chromatin architectures of rice mesophyll and endosperm, foxtail millet bundle sheath and mesophyll, and maize bundle sheath, mesophyll and endosperm tissues. We found that their global A/B compartment partitions are stable across tissues, while local A/B compartment has tissue-specific dynamic associated with differential gene expression. Plant domains are largely stable across tissues, while new domain border formations are often associated with transcriptional activation in the region. Genes inside plant domains are not conserved across species, and lack significant co-expression behavior unlike those in mammalian TADs. Although we only observed chromatin loops between gene islands in the large genomes, the maize loop gene pairs' syntenic orthologs have shorter physical distances in small genome monocots, suggesting that loops instead of domains might have conserved biological function. Our study showed that plants' chromatin features might not have conserved biological functions as the mammalian ones.

Micropapillary pattern in pure mucinous carcinoma of the breast - does it matter or not?

AIMS: Pure mucinous carcinoma (PMC) of the breast is a low-grade cancer. In this study, we aimed to elucidate the prognostic significance of micropapillary structures in breast PMC. METHODS AND RESULTS: Seventy-five patients with breast PMC were recruited. All haematoxylin and eosin (H&)-stained slides were reviewed, and clinicopathological features including age, tumour size, growth pattern, nuclear grade, histological grade, lymph node (LN) status, immunohistochemistry (IHC) staining of hormone receptor, human epidermal growth factor receptor 2 (HER2) expression and Ki-67 proliferation index were analysed. Fluorescence in-situ hybridisation (FISH) was used to verify the amplification of the HER2 gene in IHC 2+ cases. Seventy-one cases of PMC were followed-up from 18 to 110 months (median = 68 months). All PMC patients were female, aged 31-83 years (median = 57 years). All PMCs were nuclear grades 1 or 2. Oestrogen receptor was positive in all cases (100%) and progesterone receptor was positive in 68 cases (90.7%). There was no 3+ staining or gene amplification of HER2. Four patients had axillary LN metastasis (5.7%). Micropapillae were observed in 60 cases (80%) with varied percentages, and divided into five groups: 0%, <20%, 20-49%, 50-89% and ≥90%, with 15 (20%), 15 (20%), 17 (22.7%), 17 (22.7%) and 11 (14.7%) cases in each. Follow-up results showed that neither recurrence nor distant metastasis occurred in all PMCs. Statistical analysis revealed that only larger tumour size was correlated significantly with LN metastasis (P < 0.05). CONCLUSIONS: The presence of nuclear grades 1 or 2 micropapillae, irrespective of the percentage, had no significant relationship with LN metastasis and patients' survival of PMC.

The ovarian estrogen synthesis function was impaired in Y123F mouse and partly restored by exogenous FSH supplement.

BACKGROUND: LepR tyrosine site mutation mice (Y123F) exhibit decreased serum E2 levels, immature reproductive organs, infertility as well as metabolic abnormalities. Although the actions of leptin and lepR in the control of reproductive function are thought to be exerted mainly via the hypothalamic-pituitary-gonadal axis, relatively less is known regarding their local effects on the peripheral ovary, especially on steroid hormone synthesis. Meanwhile, whether the decreased fertility of Y123F mouse could be restored by gonadotropin has not been clear yet. METHODS: The serum levels of E2, P4, FSH, LH, T and leptin of Y123F and WT mice at the age of 12 weeks were measured by enzyme-linked immunosorbent assay. Immunohistochemistry was used to compare the distribution of hormone synthases (STAR, CYP11A1, CYP19A1, HSD17B7) and FSHR in adult mouse ovaries of two genotypes. Western blot and real-time PCR were used to detect the expression levels of four ovarian hormone synthases and JAK2-STAT3 / STAT5 signaling pathway in 4 and 12 weeks old mice, as well as the effects of exogenous hFSH stimulation on hormone synthases and FSHR. RESULTS: Compared with WT mice, the serum levels of FSH, LH and E2 in 12-week-old Y123F mice were significantly decreased; T and leptin levels were significantly increased; but there was no significant difference of serum P4 levels. STAR, CYP11A1, HSD17B7 expression levels and the phosphorylation levels of JAK2 and STAT3 were significantly decreased in adult Y123F mice, while the expression of CYP19A1 and phospho-STAT5 were significantly increased. No significant differences were found between 4-week-old Y123F and WT mice. After exogenous hFSH stimulation, E2 levels and expression of CYP19A1 and HSD17B7 were significantly higher than that in the non-stimulated state, but significant differences still existed between Y123F and WT genotype mice under the same condition. CONCLUSIONS: Abnormal sex hormone levels of Y123F mice were due to not only decreased gonadotropin levels in the central nervous system, but also ovarian hormone synthase abnormalities in the peripheral gonads. Both FSH signaling pathway and JAK2-STAT3/STAT5 signaling pathway were involved in regulation of ovarian hormone synthases expression. Exogenous FSH just partly improved the blood E2 levels and ovarian hormone synthase expression.

[Small cell carcinoma of ovary, hypercalcemic type: analysis of clinicopathologic characteristics and the diagnostic utility of loss expression of SMARCA4 protein].

OBJECTIVE: To study the clinicopathologic features of small cell carcinoma of ovary, hypercalcemic type (SCCOHT) and to evaluate the diagnostic significance of loss of SMARCA4 expression. METHODS: The clinicopathologic characteristics of 5 cases of SCCOHT were reviewed. The expression of SMARCA4 protein was detected by immunohistochemistry in the cases of SCCOHT and 240 cases of other primary malignant tumors of ovary and peritoneum. RESULTS: The mean and medium age of these patients was 30 years and 28 years, respectively. The presenting symptoms included abdominal pain, distention and a pelvic mass. Hypercalcemia was found in 3 patients. The maximum diameter of tumors ranged from 13.5 to 22.0 cm. Extraovarian spread was demonstrated in all of the patients on presentation. Histologically, the tumors were composed of closely packed small round cells with scanty cytoplasm, hyperchromatic nuclei and irregular chromatin clumps. The tumor cells grew in sheets, nests, cords or trabecular pattern. Follicle-like spaces were observed in 4 cases. Three of the tumors contained large cells with abundant eosinophilic cytoplasm. Spindle cell morphology was found in 1 case. There were 2 cases with myxoid or hyaline stroma. Four out of five of SCCOHT cases showed loss of SMARCA4 protein while only 6.3% (15/240) of the other primary malignant tumors of ovary and peritoneum , including undifferentiated carcinoma (1/5), high-grade serous carcinoma (4.6%, 5/109), endometrioid carcinoma (7.7%, 2/26), clear cell carcinoma (1/9), mucinous carcinoma (1/5), mixed carcinoma (4.9%, 3/61), carcinosarcoma (1/9) and high-grade serous carcinoma of peritoneum (1/9), were negative. CONCLUSIONS: SCCOHT is a rare malignant tumor and often misdiagnosed as other types of ovarian small cell tumor. Loss expression of SMARCA4 protein is characteristic and facilitates the diagnosis and differential diagnosis of SCCOHT.

[Metastases to the breast from non-mammary malignancies: a clinicopathologic study of 28 cases].

OBJECTIVE: To investigate the clinicopathologic characteristics and differential diagnosis of the metastases to the breast from non-mammary malignancies. METHODS: Twenty-eight cases were collected from 2004 to 2012;microscopic pathologic examinations and immunohistochemistry (EnVision method) were performed. RESULTS: (1) All except one patients were female, ranging from 16 to 77 years old (average 45.8 years). Twenty-six (92.9%) patients initially presented with the primary site lesions; while the other two (7.1%) patients initially presented with breast lesions. The mean interval from primary diagnosis to detection of metastatic breast lesions was 32 months (0-228 months). Fifteen patients (53.6%) had other metastases detected simultaneously or preceded the breast lesions. (2) Macroscopically, all the tumors were relatively circumscribed, with a mean diameter of 4.0 cm (0.6-12.0 cm). The histological types of the corresponding primary tumors were as follows: eight (28.6%) cases from lung adenocarcinoma, five (17.8%) from high-grade ovarian serous carcinoma, three (10.7%) from gastric adenocarcinoma, two (7.1%) from rectal adenocarcinoma, one (3.6%) from pancreatic neuroendocrine carcinoma, one (3.6%) from prostatic carcinoma, four (14.3%) from melanoma, and four (14.3%) from mesenchymal malignant tumors (three rhabdomyosarcomas and one epithelioid malignant peripheral nerve sheath tumor, MPNST). (3) Histologically, the metastatic tumors showed the morphologic characteristics of the primary tumors. Lymph-vascular invasion was observed in 19 cases. Immunohistochemical features of metastatic tumors were consistent with the primary tumors. Molecular markers for breast such as GCDFP15 and mammaglobin were negative. Metastatic tumors from lung adenocarcinoma expressed TTF-1 (8/8). Ovarian serous carcinoma metastases were positive for PAX8 (5/5) and WT1 (4/5). Gastric adenocarcinoma metastases were positive for CDX2 (3/3) and villin (1/3). Rectal adenocarcinoma metastases were positive for CDX2 (2/2). Pancreatic neuroendocrine tumor metastasis was positive for Syn and CgA (both 1/1). Prostate carcinoma metastasis was positive for AR, PSA and P504S (all 1/1). Melanoma metastases were positive for HMB45 (2/3) and S-100 protein (3/3). Rhabdomyosarcoma metastases were positive for vimentin, desmin and myoD1 (all 3/3). MPNST metastasis was positive for S-100 protein (1/1). (4) Follow-up data was available in 17 patients, with median follow-up time 54 months. The median survival from diagnosis to breast metastasis was 24 months.Seven of 17 patients died. CONCLUSIONS: Metastases to the breast from non-mammary malignancies are rare and show pathologic features of primary tumors. It is usually presumed to be a primary breast carcinoma. Histopathologic features and clinical history in conjunction with the immunohistochemical results should be considered in differentiating a secondary mass from a primary breast carcinoma.

Investigating the cis-Regulatory Basis of C3 and C4 Photosynthesis in Grasses at Single-Cell Resolution.

While considerable knowledge exists about the enzymes pivotal for C4 photosynthesis, much less is known about the cis-regulation important for specifying their expression in distinct cell types. Here, we use single-cell-indexed ATAC-seq to identify cell-type-specific accessible chromatin regions (ACRs) associated with C4 enzymes for five different grass species. This study spans four C4 species, covering three distinct photosynthetic subtypes: Zea mays and Sorghum bicolor (NADP-ME), Panicum miliaceum (NAD-ME), Urochloa fusca (PEPCK), along with the C3 outgroup Oryza sativa. We studied the cis-regulatory landscape of enzymes essential across all C4 species and those unique to C4 subtypes, measuring cell-type-specific biases for C4 enzymes using chromatin accessibility data. Integrating these data with phylogenetics revealed diverse co-option of gene family members between species, showcasing the various paths of C4 evolution. Besides promoter proximal ACRs, we found that, on average, C4 genes have two to three distal cell-type-specific ACRs, highlighting the complexity and divergent nature of C4 evolution. Examining the evolutionary history of these cell-type-specific ACRs revealed a spectrum of conserved and novel ACRs, even among closely related species, indicating ongoing evolution of cis-regulation at these C4 loci. This study illuminates the dynamic and complex nature of CRE evolution in C4 photosynthesis, particularly highlighting the intricate cis-regulatory evolution of key loci. Our findings offer a valuable resource for future investigations, potentially aiding in the optimization of C3 crop performance under changing climatic conditions.

Autophagy plays a pro-apoptotic role in arsenic trioxide-induced cell death of liver cancer.

OBJECTIVE: The effects of arsenic trioxide (As2O3) on hepatocellular carcinoma have been documented widely. Autophagy plays dual roles in the survival and death of cancer cells. Therefore, we investigated the exact role of autophagy in As2O3-induced apoptosis in liver cancer cells. METHODS: The viability of hepatoma cells was determined using the MTT assay with or without fetal bovine serum. The rate of apoptosis in liver cancer cells treated with As2O3 was evaluated using flow cytometry, Hoechst 33258 staining, and TUNEL assays. The rate of autophagy among liver cancer cells treated with As2O3 was detected using immunofluorescence, Western blot assay and transmission electron microscopy. RESULTS: Upon treatment with As2O3, the viability of HepG2 and SMMC-7721 cells was decreased in a time- and dose-dependent manner. The apoptosis rates of both liver cancer cell lines increased with the concentration of As2O3, as shown by flow cytometry. Apoptosis in liver cancer cells treated with As2O3 was also shown by the activation of the caspase cascade and the regulation of Bcl-2/Bax expression. Furthermore, As2O3 treatment induced autophagy in liver cancer cells; this finding was supported by Western blot, immunofluorescence of LC3-II and beclin 1, and transmission electron microscopy. In liver cancer cells, As2O3 inhibited the phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/AKT/mTOR) signal pathway that plays a vital role in both apoptosis and autophagy. The PI3K activator SC-79 partially reversed As2O3-induced autophagy and apoptosis. Furthermore, inhibiting autophagy with 3-methyladenine partially reversed the negative effects of As2O3 on cell viability. Serum starvation increased autophagy and amplified the effect of As2O3 on cell death. CONCLUSION: As2O3 induces apoptosis and autophagy in liver cancer cells. Autophagy induced by As2O3 may have a proapoptotic effect that helps to reduce the viability of liver cancer cells. This study provides novel insights into the effects of As2O3 against liver cancer. Please cite this article as: Deng ZT, Liang SF, Huang GK, Wang YQ, Tu XY, Zhang YN, Li S, Liu T, Cheng BB. Autophagy plays a pro-apoptotic role in arsenic trioxide-induced cell death of liver cancer. J Integr Med. 2024; 22(3): 295-302.

Short-term effects of cupping and scraping therapy for chronic nonspecific low-back pain: A prospective, multicenter randomized trial.

BACKGROUND: As one of the most common musculoskeletal ailments, chronic nonspecific low-back pain (CNLBP) causes persistent disability and substantial medical expenses. Epidemiological evidence shows that the incidence rate of CNLBP in young and middle-aged people who are demanded rapidly recovery and social contribution is rising. Recent guidelines indicate a reduced role for medicines in the management of CNLBP. OBJECTIVE: The present study investigates the short-term effects of cupping and scraping therapy using a medicated balm, compared to nonsteroidal anti-inflammatory drug (NSAID) with a capsaicin plaster, in the treatment of CNLBP. DESIGN, SETTING, PARTICIPANTS AND INTERVENTIONS: We designed a prospective multicenter randomized clinical trial enrolling patients from January 1, 2022 to December 31, 2022. A total of 156 patients with CNLBP were randomized into two parallel groups. Diclofenac sodium-sustained release tablets were administered orally to participants in the control group for one week while a capsaicin plaster was applied externally. Patients in the test group were treated with cupping and scraping using a medical device and medicated balm. MAIN OUTCOME MEASURES: Primary outcome was pain recorded using the visual analogue scale (VAS). Two secondary outcomes were recorded using the Japanese Orthopedic Association low-back pain scale (JOA) and the traditional Chinese medicine (TCM) syndrome integral scale (TCMS) as assessment tools. RESULTS: Between baseline and postintervention, all changes in outcome metric scales were statistically significant (P < 0.001). Compared to the control group, patients in the test group had a significantly greater treatment effect in all outcome variables, as indicated by lower VAS and TCMS scores and higher JOA scores, after the one-week intervention period (P < 0.001). Further, according to the findings of multivariate linear regression analysis, the participants' pain (VAS score) was related to their marital status, age, smoking habits and body mass index. No adverse reactions were reported for any participants in this trial. CONCLUSION: The effectiveness of TCM combined with the new physiotherapy tool is superior to that of NSAID combined with topical plasters, regarding to pain intensity, TCM symptoms and quality of life. The TCM plus physiotherapy also showed more stable and long-lasting therapeutic effects. TRIAL REGISTRATION: This study was registered at Chinese Clinical Trial Registry (ChiCTR2200055655). Please cite this article as: He JY, Tu XY, Yin ZF, Mu H, Luo MJ, Chen XY, Cai WB, Zhao X, Peng C, Fang FF, Lü C, Li B. Short-term effects of cupping and scraping therapy for chronic nonspecific low-back pain: A prospective, multicenter randomized trial. J Integr Med. 2024; 22(1): 39-45.

Evolution of plant cell-type-specific cis-regulatory elements.

Cis-regulatory elements (CREs) are critical in regulating gene expression, and yet understanding of CRE evolution remains challenging. Here, we constructed a comprehensive single-cell atlas of chromatin accessibility in Oryza sativa, integrating data from 103,911 nuclei representing 126 discrete cell states across nine distinct organs. We used comparative genomics to compare cell-type resolved chromatin accessibility between O. sativa and 57,552 nuclei from four additional grass species (Zea mays, Sorghum bicolor, Panicum miliaceum, and Urochloa fusca). Accessible chromatin regions (ACRs) had different levels of conservation depending on the degree of cell-type specificity. We found a complex relationship between ACRs with conserved noncoding sequences, cell-type specificity, conservation, and tissue-specific switching. Additionally, we found that epidermal ACRs were less conserved compared to other cell types, potentially indicating that more rapid regulatory evolution has occurred in the L1-derived epidermal layer of these species. Finally, we identified and characterized a conserved subset of ACRs that overlapped the repressive histone modification H3K27me3, implicating them as potentially silencer-like CREs maintained by evolution. Collectively, this comparative genomics approach highlights the dynamics of plant cell-type-specific CRE evolution.

Response to anti-HER2 neoadjuvant chemotherapy in HER2-positive invasive breast cancers with different HER2 FISH patterns.

AIMS: Human epidermal growth factor receptor 2 (HER2)-positive patients with breast cancer may have different HER2/CEP17 ratios and HER2 copy numbers, with inconsistent responses to anti-HER2 neoadjuvant chemotherapy (NACT). Our study aimed to explore the relationship between different HER2 fluorescence in situ hybridisation (FISH) patterns in HER2-positive patients with breast cancer and responses to anti-HER2 NACT. METHODS: 527 patients with HER2-positive invasive breast cancer who received anti-HER2 NACT from 2015 to 2022 were included and divided into three groups by FISH results, namely group A: HER2/CEP17<2.0 and HER2 copy numbers ≥6.0, HER2 immunohistochemistry 2/3+; group B: HER2/CEP17≥2.0 and HER2 copy numbers ≥4.0 and <6.0; group C: HER2/CEP17≥2.0 and HER2 copy numbers ≥6.0. We compared clinicopathological characteristics and pathological complete response (pCR) rates of different groups. RESULTS: According to HER2 FISH results, 12 patients (2.3%, 12/527) were in group A, 40 (7.6%, 40/527) were in group B and 475 (90.1%, 475/527) were in group C. The pCR rate was the lowest in group B (5.0%), while the pCR rates in group A and group C were 33.3% and 44.4%, respectively (p (group A vs. B) =0.021, p (group C vs. B) < 0.001). Both univariate and multivariate analyses revealed that HER2 FISH pattern was correlated with pCR rate (p (group C vs. B) < 0.001, p (group C vs. B) = 0.025). CONCLUSIONS: Patients with HER2/CEP17≥2.0 and HER2 copy numbers ≥4.0 and <6.0 do not benefit to the same extent from current anti-HER2 therapies as FISH-positive patients with other patterns.

A prospective study of fertility-sparing treatment with megestrol acetate following hysteroscopic curettage for well-differentiated endometrioid carcinoma and atypical hyperplasia in young women.

PURPOSE: To investigate the feasibility and efficacy of curettage with hysteroscopy followed by megestrol acetate (MA) for well-differentiated endometrioid carcinoma (EC) confined to the endometrium and for atypical hyperplasia (AH) in young women. PATIENTS AND METHODS: Fourteen patients with EC and 12 patients with AH were prospectively enrolled in this study. All of the patients received at least 12 weeks of oral MA (160 mg/day) following thorough curettage with hysteroscopy. The response was assessed histologically every 12 weeks. The primary endpoint was the complete response rate. Adverse events, pregnancy rates and recurrence rates were secondary end points. RESULTS: Twenty-one (80.8 %) patients responded to treatment. The median time to response was 12 weeks. After a median follow-up of 32 months, 6 patients had recurrences. Significantly, more patients with infertility or PCOS experienced recurrence (P = 0.040, P = 0.015). Eight patients attempted to conceive after complete response; two spontaneous conceptions and one normal delivery were achieved. No disease-related or treatment-related deaths were observed. CONCLUSIONS: Fertility-sparing treatment with MA following entirely hysteroscopic curettage is effective, demonstrating the least toxicity for rigorously selected young women with well-differentiated EC confined to the endometrium or with AH; however, close follow-up is required for the potential consequences of improper patient selection and a substantial rate of recurrence.

Intramolecular Ring Expansion of 3-Silaazetidine with Alkynes Enabled by Pd-Catalyzed Si-C Bond Activation.

An intramolecular ring expansion of in situ formed 3-silaazetidine with internal alkynes has been developed via Pd-catalyzed Si-C bond activation. The reaction gives rise to 6,5- and 6,6-fused bicyclic 1,3-azasilines, in which the silicon atom locates at the ring junction position.