Enhanced prognostic evaluation of diffuse large B-cell lymphoma: A comprehensive surveillance study incorporating Epstein-Barr virus infection status and immunohistochemical markers.

Emerging biologic subsets and new prognostic markers are significantly important for aggressive diffuse large B-cell lymphoma (DLBCL). Nevertheless, the high cost of testing limits the availability of these tests in most hospitals, thus making prognostic judgment based on basic immunohistochemical testing, whole blood Epstein-Barr virus DNA (WBEBV) surveillance and clinical features advantageous for hospitals and patients with poor medical conditions. We included 647 DLBCL patients treated in our hospital from January 2009 to March 2023. Non-germinal center B-cell like, Ki-67, and International Prognostic Index (IPI) scores were related to cMYC/B-cell lymphoma 2 (Bcl-2)-double expression. Age, Epstein-Barr virus-encoded small RNA (EBER) positivity, and IPI scores were associated with mortality. The cutoffs for differential overall survival (OS) of age, WBEBV, Bcl-2, and cMYC were 57 years, 1514 copies/mL (baseline), 5.89 × 104 copies/mL (treatment), 40%, and 55%, respectively. EBER positivity was significantly associated with a worse OS. Patients with newly defined DE (Bcl-2 ≥ 40 and cMYC > 55) had a worse prognosis than controls (p = 0.04). We found that cMYC with an optimal cutoff of 47.5 could effectively predict high-grade DLBCL with an area under the curve of 0.912, and the specificity and sensitivity were 70.7% and 100%, respectively. Our study provides valuable insights into the prognostic factors and biomarker cutoffs that influence OS in DLBCL patients, which may guide clinicians in tailoring treatment strategies and improving patient outcomes.

Ocimum basilicum L. (basil) presents pro-apoptotic activity in an Ehrlich's experimental tumor murine model.

PURPOSE: This study aimed to evaluate the therapeutic effect of an ethanol extract of Ocimum basilicum L. (EEOb) aerial parts against Ehrlich's experimental tumor (EET) in mice. METHODS: Swiss mice were divided into two groups (control and treated; n = 6). On day 21, all mice were inoculated subcutaneously with 2 × 106 (0.05 mL) EET cells in the left paw for solid tumor development. This study lasted 28 days. Treatment began 24 hours after inoculation with EET. Measurements of dorsoplantar thickness were used to assess tumor growth. The paw pad was collected for histopathological analysis and stained using the argyrophilic nucleolar organizing regions (AgNOR) technique and immunohistochemistry for proliferating cell nuclear antigen, Bcl-2 and Bax. RESULTS: The treatment of animals with EEOb at 100 mg/kg intraperitoneally was able to reduce the growth (Control = 3.7 ± 0.1 mm vs. EEOb = 5.7 ± 0.2 mm) and the number of AgNORs of solid Ehrlich tumor. The antitumor effect of EEOb was associated with the induction of apoptosis of tumoral cell, as suggested by the reduction of the content of Bcl-2 induced by extract. CONCLUSIONS: The study demonstrated that daily administration of EEOb is able to reduce the growth of EET by induce apoptosis of tumoral cells.

Assessment and comparative study of diosgenin doses in alleviating experimental periodontitis.

BACKGROUND: This study was performed to determine the therapeutic effects of diosgenin (DG) which is a steroidal saponin, administered at different doses on alveolar bone loss (ABL) in rats with experimental periodontitis using immunohistochemical and cone-beam computed tomography (CBCT). METHODS: Thirty-two male Wistar rats divided into four equal groups: control (non-ligated), periodontitis (P), DG-48, and DG-96. Sutures were placed at the gingival margin of the lower first molars to induce experimental periodontitis. Then, 48 and 96 mg/kg of DG was administered to the study groups by oral gavage for 29 days. At day 30, the animals were sacrificed and ABL was determined via CBCT. The expression patterns of osteocalcin (OCN), alkaline phosphatase (ALP), type I collagen (Col-1), B cell lymphoma 2 (Bcl 2), Bcl 2-associated X protein (Bax), bone morphogenetic protein 2 (BMP-2), and receptor activator of NF κB ligand (RANKL) were examined immunohistochemically. RESULTS: Histopathologic examination showed all features of the advanced lesion in the P group. DG use decreased all these pathologic changes. It was observed that periodontitis pathology decreased as the dose increased. DG treatment increased the ALP, OCN, Bcl 2, Col-1, and BMP-2 levels in a dose-dependent manner, compared with the P group (p < 0.05). DG decreased the expression of RANKL and Bax in a dose-dependent manner (p < 0.05). ABL was significantly lower in the DG-48 and DG-96 groups than in the P group (p < 0.05). CONCLUSION: Collectively, our findings suggest that DG administration protects rats from periodontal tissue damage with a dose-dependent manner, provides an increase in markers of bone formation, decreases in Bax/Bcl-2 ratio and osteoclast activation.

MDS and AML show elevated fractions of CD34-positive blast cell populations with a high anti-apoptotic versus proliferation ratio.

This study investigates the intertwined processes of (anti-)apoptosis and cell proliferation in myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). Utilizing antibodies to Bcl-2 and Ki-67, the CD34-positive blast cell compartments in bone marrow aspirates from 50 non-malignant cases, 25 MDS patients, and 25 AML patients were analyzed for their anti-apoptotic and proliferative cell fractions through ten-color flow cytometry. MDS patients exhibited a significantly increased anti-apoptotic (p=0.0014) and reduced proliferative cell fraction (p=0.0030) in their blast cell population as compared to non-malignant cases. AML patients showed an even more exacerbated trend than MDS patients. The resulting Bcl-2:Ki-67 cell fraction ratios in MDS and AML were significantly increased as compared to the non-malignant cases (p=0.0004 and p<0.0001, respectively). AML patients displayed, however, a high degree of variability in their anti-apoptotic and proliferation index, attributed to heterogeneity in maturation stage and severity of the disease at diagnosis. Using double-labeling for Bcl-2 and Ki-67 it could be shown that besides blast cells with a mutually exclusive Ki-67 and Bcl-2 expression, also blast cells concurrently exhibiting anti-apoptotic and proliferative marker expression were found. Integrating these two dynamic markers into MDS and AML diagnostic workups may enable informed conclusions about their biological behavior, facilitating individualized therapy decisions for patients.

A novel triad for the diagnosis of endometriosis, the short anogenital distance combines with high endometrial BCL2 and low endometrial FASL.

OBJECTIVE: To investigate the anogenital distance from the upper verge of the anus to the posterior fourchette (AGDAF), FASL, and BCL2 combination as a reliable and non-invasive tool for the diagnosis of endometriosis. METHODS: This study included 100 women with endometriosis and 50 women without endometriosis as the control group. All cases underwent history taking, body mass index (BMI) measurement, AGD measurement, and FASL and BCL2 immunohistochemical staining of the eutopic endometrial tissue. RESULTS: This study included 150 women divided into endometriosis and control groups. Endometriosis cases significantly had shorter AGDAF, 22.9 ± 2.6 mm, compared with the control group, 27.3 ± 3.5 mm (P < 0.001). Lower FASL and higher BCL2 expression were associated with endometriosis (P < 0.001). The combined measurement of AGDAF (cut-off point 24.55 mm) with FASL and BCL2 was associated with endometriosis (P < 0.001). The combined diagnostic sensitivity, specificity, positive predictive value, and negative predictive value of AGDAF, FASL, and BCL2 were 83%, 78%, 87.3%, and 69.6%, respectively. The area under the curve was greater for AGDAF, FASL, and BCL2 in combination than for individual measurements. CONCLUSION: Combining short AGDAF with high BCL2 and low FASL is a highly sensitive, non-invasive diagnostic tool for endometriosis.

Clinical relevance of MYC/BCL2 expression and cell of origin in patients with diffuse large b-cell lymphoma treated with autologous transplant.

Dual expression of MYC and BCL2 proteins (double-expressor lymphoma [DEL]) as well as cell of origin (COO) are important prognostic factors in patients with diffuse large B-cell lymphoma (DLBCL) after conventional chemotherapy. We studied the prognostic impact of DEL and COO in patients with relapsed DLBCL treated with autologous stem cell transplant (ASCT). Three-hundred and three patients with stored tissue samples were identified. Classification was successful in 267 patients: 161 (60%) were DEL/non-double hit (DHL), 98 (37%) were non-DEL/non-DHL, and 8 (3%) were DEL/DHL. Compared to non-DEL/non-DHL, DEL/DHL had worse overall survival while DEL/non-DHL did not significantly differ in overall survival. On multivariable analysis, DEL/DHL, age >60 years, and >2 prior therapies, but not COO, were important prognostic factors for overall survival. When we explored the interaction of COO and BCL2 expression, patients with germinal center B-cell (GCB)/BCL2 (+) had inferior progression-free survival (PFS) compared to GCB/BCL2 (-) patients (HR, 4.97; P = 0.027). We conclude that the DEL/non-DHL and non-DEL/non-DHL subtypes of DLBCL have similar survival after ASCT. The negative impact of GCB/BCL2 (+) on PFS warrants future trials targeting BCL2 after ASCT. The inferior outcomes in DEL/DHL need to be verified in a larger number of patients.

Genetic Profiling of Diffuse Large B-Cell Lymphoma: A Comparison Between Double-Expressor Lymphoma and Non-Double-Expressor Lymphoma.

INTRODUCTION: Data are limited regarding the genetic profiling of diffuse large B-cell lymphoma (DLBCL) with double expression of MYC and BCL2 proteins without underlying rearrangements (double-expressor lymphoma [DEL]). This study aimed to describe the genetic profiling and determine the prognostic significance in patients with DEL and in those with non-DEL. METHODS: Capture-based targeted sequencing was performed on 244 patients with de novo DLBCL, not otherwise specified. Immunohistochemistry staining was performed for evaluating the MYC and BCL2 expression. RESULTS: Among 244 patients, 46 patients had DEL, and 198 had non-DEL. KMT2D, CD58, EP300, PRDM1, TNFAIP3 and BCL2 gain or amplification (BCL2GA/AMP) were significantly more frequently altered in the DEL group. Alterations in the BCR/TLR (p = 0.021), B-cell development and differentiation (p = 0.004), and NF-κB (p = 0.034) pathways occurred more frequently in patients with DEL. Thirty-seven DEL patients and 132 non-DEL patients were included for survival analyses. DEL was not significantly associated with progression-free survival (PFS) (p = 0.60) and overall survival (OS) (p = 0.49). In DEL patients, after adjusting for the International Prognostic Index, BCL2 alteration (HR 2.516, 95% CI 1.027-6.161; p = 0.044) remained an independent predictor of inferior PFS. BCL2GA/AMP also predicted poor PFS, but with marginal statistical significance (HR 2.489, 95% CI 0.995-6.224; p = 0.051). CONCLUSION: There was difference in profiling of altered genes and signaling pathways between the DEL group and the non-DEL group. The presence of DEL alone should not be considered as an adverse prognostic indicator, and BCL2 alteration could define a subset of patients with poor prognosis within DEL.

Association between redundant endometrium and endometrial polyps: a pilot study.

BACKGROUND: The aim of this study was to explore the organic features of redundant endometrium (RE), we examined the expression of different endometrial hormone receptors, oncogenes, and cell replication markers, in normal endometrium (NE), endometrial polyps (EP) and RE specimens. METHODS: This was an experimental study examining endometrial tissue expression of estrogen receptors (ER1 and 2), progesterone receptors (PR-A+B), androgen receptor (AR), insulin receptor (Insulin-R), insulin-like growth factor receptor 1 (IGFR-1), thyroid hormone receptor (TH-RB), B-cell lymphoma 2 (Bcl-2), Ki67, HOXA10, in women with NE, EP and RE, of women undergoing hysteroscopy for benign gynecologic pathology. Specimens were separated in 3 groups: NE, EP, RE. Endometrial samples were processed for real-time RT-PCR analyses. Main outcome measure was tissue expression of the markers in the three groups. RESULTS: Of the 16 patients, 2 had NE, 8 had RE, 5 had EP, 1 had both, RE and EP. Compared to NE, RE and EP showed significantly increased Bcl-2, Insulin-R, ER-ß, PR-A+B, and TRB expression (P<0.044), with EP showing significantly increased PR-A+B, compared to RE (3.29±0.47 fg/µg RNA versus 1.86±0.34 fg/µg RNA; P=0.023). The other markers were not significantly different across the three groups: Ki67 appeared non-significantly decreased, while HOXA10, IGF-R1, AR, and ER-α, were non-significantly increased. CONCLUSIONS: RE showed biochemical characteristics different from NE. Similar to endometrial polyps, RE showed enhanced cell differentiation, but not cell replication. These changes in RE could be detrimental for embryo implantation and should be of consideration in women undergoing fertility treatments.

Prognostic value of Bcl2 and Bcl6 in primary cutaneous large B-cell lymphoma: A systematic review and meta-analysis.

AIMS: To assess the prognostic value of Bcl2 and Bcl6 in primary cutaneous diffuse large B-cell lymphoma (pcDLBCL), through a systematic review and meta-analysis. METHODS: Electronic databases were searched from their inception to April 2021 for studies reporting Bcl2 and Bcl6 expression and survival outcomes in pcDLBCL series. Kaplan-Meier and Cox regression survival analyses with hazard ratio calculation were performed for overall survival (OS), with a significant p-value< 0.05. RESULTS: Eight studies with 148 patients were included. OS was significantly decreased in Bcl2-pos itive pcDLBCLs (5-year OS= 52.9 ± 5.2%) compared to Bcl2 negative pcDLBCLs (5-year OS= 86.6 ± 7.2%), with a HR of 4.615 (95% CI, 1.827-11.657; p = 0.001); no significant difference in OS was found between Bcl6-positive pcDLBCLs (5-year OS= 61.3 ± 6.5%) and Bcl6-negative pcDLBCLs (5-year OS= 56.8 ± 7.2%), with a HR of 0.789 (95% CI, 0.462-1.350; p = 0.388). CONCLUSIONS: In pcDLBCL, Bcl2 expression is a strong unfavourable prognostic marker; Bcl6 does not seem to be associated with survival instead. Further studies are necessary in this field.

Single-Cell Visualization of Monogenic RNA G-quadruplex and Occupied G-quadruplex Ratio through a Module-Assembled Multifunctional Probes Assay (MAMPA).

G-quadruplexes (G4s), non-canonical nucleic acid secondary structure, regulate many biological functions and are considered potential molecular targets for cancer therapeutics. However, due to the lack of analytical methods, the regulating mechanism of monogenic G4s is still unclear. Here, we developed a Module Assembled Multifunctional Probes Assay (MAMPA) for visualizing endogenous G4s in individual genes in single cells. Two modular probes separately recognize G4 structures and the adjacent RNA sequences, and the module assembly enables imaging of G4s in an individual RNA with high specificity. Through imaging G4s in several individual genes, we found that G4s were steadily occupied by G4 Binding Proteins (G4BPs) in various mRNAs in every cell line and defined "Occupied G4 Ratio". We demonstrated MAMPA was suitable for most experimental situations and found that Occupied G4 Ratios had the potential to become a new parameter for the study of G4s in living cells.

Clinicopathologic and Prognostic Significance of Bruton's Tyrosine Kinase Expression in Diffuse Large B-Cell Lymphoma.

BACKGROUND/AIM: Bruton's tyrosine kinase (BTK)-mediated B-cell-receptor signaling drives lymphomagenesis of diffuse large B-cell lymphoma (DLBCL). We investigated the clinicopathological significance of BTK positivity in DLBCL according to known molecules related to resistance to BTK inhibitors [BCL2 apoptosis regulator (BCL2)/MYC proto-oncogene, bHLH transcription factor (MYC)]. PATIENTS AND METHODS: We evaluated BTK expression immunohistochemically in 106 DLBCLs considering their BCL2/MYC status. RESULTS: Considering the whole cohort, BTK was expressed in 65.1%, including 70.4% (50/71) of non-germinal center B-cell-like (non-GCB) subtype; BCL2 expression was detected in 60.4%, MYC expression in 15.1%, MYC translocation in 4.2% (4/96) and MYC gain/amplification in 7.6% (8/105). Overall and in the non-GCB cohort, BTK positively correlated with high international prognostic index (both p=0.005) and stage (p=0.006 and p=0.002), and with BCL2 intensity (p=0.005 and p=0.026, respectively); MYC gain/amplification total cohort (p=0.038). Moreover, high risk, defined as co-expression of BTK and either or both BCL2/MYC, independently predicted shorter progression-free survival in patients treated with rituximab plus cyclophosphamide, doxorubicin, vincristine and prednisone (R-CHOP) (all R-CHOP-treated patients: hazard ratio=2.565, p=0.044; R-CHOP-treated non-GCB subgroup: HR=3.833, p=0.019). CONCLUSION: BTK expression may be utilized to stratify risk in patients with DLBCL.

Cell death and pathological findings of the spleen in COVID-19 patients.

The coronavirus disease 2019(COVID-19) is recognized as systemic inflammatory response syndrome. It was demonstrated that a rapid increase of cytokines in the serum of COVID-19 patients is associated with the severity of disease. However, the mechanisms of the cytokine release are not clear. By using immunofluorescence staining we found that the number of CD11b positive immune cells including macrophages in the spleens of died COVID-19 patients, was significantly higher than that of the control patients. The incidence of apoptosis as measured by two apoptotic markers, TUNEL and cleaved caspase-3, in COVID-19 patients' spleen cells is higher than that in control patients. By double immunostaining CD11b or CD68 and SARS-CoV-2 spike protein, it was found that up to 67% of these immune cells were positive for spike protein, suggesting that viral infection might be associated with apoptosis in these cells. Besides, we also stained the autophagy-related molecules (p-Akt、p62 and BCL-2) in spleen tissues, the results showed that the number of positive cells was significantly higher in COVID-19 group. And compared with non-COVID-19 patients, autophagy may be inhibited in COVID-19 patients. Our research suggest that SARS-CoV-2 may result in a higher rate of apoptosis and a lower rate of autophagy of immune cells in the spleen of COVID-19 patients. These discoveries may increase our understanding of the pathogenesis of COVID-19.

Placental apoptotic markers are associated with placental morphometry.

INTRODUCTION: Preeclampsia is a hypertensive disorder affecting both mother and the fetus and is a major cause of maternal and neonatal morbidity and mortality. Abnormal placentation is a common feature in preeclampsia that contributes to placental dysfunction. It is likely that increased homocysteine and oxidative stress influence apoptosis in preeclampsia. Increased placental apoptosis may aggravate the symptoms of preeclampsia through disruption of the placental structure. The current study aims to examine the association between various placental apoptotic markers with placental dimensions and maternal and neonatal characteristics in women with preeclampsia. METHODS: A total of 80 pregnant women [preeclampsia (n = 40); normotensive control (n = 40)] were included in the study. Placental characteristics such as its major axis, minor axis, breadth, thickness (at centre, cord insertion and periphery) and trimmed placental weight were recorded.Placental protein levels of caspase-3, caspase-8, BAX and Bcl-2 were estimated by ELISA and gene expression were examined by real time quantitative PCR. RESULT: Protein levels of proapoptotic markers such as caspase-8 and 3 were higher (p < 0.01) in the preeclampsia group compared to control whereas, the level of antiapoptotic marker Bcl-2 (p < 0.05) was lower in the preeclampsia group. Caspase-3 and Bcl-2 protein levels were negatively associated with thickness of placenta at cord insertion (p < 0.01). Protein levels of caspase-8 and caspase-3 were positively associated with placental MDA levels (p < 0.01). Caspase-8 was negatively associated with baby length (p = 0.055). DISCUSSION: This study demonstrates the association of various apoptotic markers with oxidative stress and placental dimensions.

Effects of hyperbaric oxygen pretreatment on brain antioxidant capacity in rats with decompression sickness.

OBJECTIVE: Decompression sickness (DCS) causes serious brain hypoxic-ischemic injury. This experiment was designed to observe whether hyperbaric oxygen (HBO2) pretreatment played a neuroprotective effect in decompression sickness rat models and to explore the mechanism of protective effects. METHODS: Sprague-Dawley (SD) male rats were pretreated with HBO2 and then underwent decompression to establish the DCS rat model. Antioxidant capacities were evaluated by detecting peroxides (GPx), superoxide dismutase (SOD), catalase (CAT) activity and malondialdehyde (MDA) content in brains. The levels of metal elements manganese (Mn), zinc (Zn), iron (Fe) and magnesium (Mg) in brain tissues were assessed by flame atomic absorption spectrometry. Necrosis and apoptosis of neurons were assessed by H-E staining and immunohistochemical staining. RESULTS: HBO2 pretreatment reduced the degree of necrosis and apoptosis in brain tissues of decompression sickness rat models. In addition, HBO2 pretreatment increased GPx, SOD and CAT activities and reduced MDA accumulation. It also increased the content of Mn, Zn, Fe and Mg in brain tissue, which are all related to free radical metabolism. CONCLUSION: These results suggested that HBO2 pretreatment has protective effects on brain injury of rats with decompression sickness. The mechanism of the protective effects may be related to reducing oxidative damage by affecting metal elements in vivo.

Demethoxycurcumin Suppresses Human Brain Glioblastoma Multiforme GBM 8401 Cell Xenograft Tumor in Nude Mice In Vivo.

Demethoxycurcumin (DMC), a derivate of curcumin, has been shown to induce apoptotic cell death in human glioblastoma multiforme GBM 8401 cells via cell cycle arrest and induction of cell apoptosis. However, there is no report showing DMC suppresses glioblastoma multiforme cells in vivo. In the present study, we investigated the effects of DMC on GBM8401 cells in vivo. At first, we established a luciferase-expressing stable clone named GBM 8401/luc2. Second, mice were inoculated subcutaneously with GBM 8401/luc2 cells to generate a xenograft tumor mice model. After inoculation, tumor volume reached 100-120 mm3, and all mice were randomly divided into three groups: Group I was treated with 110 µL phosphate-buffered solution (PBS) containing 0.1% dimethyl sulfoxide, Group II with 30 mg/kg of DMC, and Group III with 60 mg/kg of DMC. Mice from each group were given the oral treatment of DMC by gavage for 21 days. The body weight and tumor volume were recorded every 3 days. DMC significantly decreased the tumor volumes, and 60 mg/kg treatment showed a higher decrease in tumor volumes than that of 30 mg/kg, However, DMC did not affect the body weights. The photons emitted from mice tumors were detected with Xenogen IVIS imaging system, DMC at both doses decreased the total photon flux and 60 mg/kg treatment of DMC has low total photon flux than that of 30 mg/kg. The tumor volumes and weights in 60 mg/kg treatment of DMC were lower than that of 30 mg/kg. Immunohistochemical analysis was used to measure protein expression of tumors and results showed that DMC treatment led to lightly staining with anti-Bcl-2 and -XIAP and 60 mg/kg treatment of DMC has lighter staining with anti-Bcl-2 and -XIAP than that of 30 mg/kg. The higher dose (60 mg/kg) of DMC has higher signals of cleaved-caspase-3 than that of the lower dose (30 mg/kg). Furthermore, the hematoxylin and eosin (H&E) staining of liver tissues showed no significant difference between DMC-treated and control-groups. Overall, these observations showed that DMC suppressed tumor properties in vivo and DMC may be used against human glioblastoma multiforme in the future.

Combination of biomarkers for neoadjuvant systemic chemotherapy before cystectomy in patients with urinary bladder cancer.

Clinical utility of cisplatin based neoadjuvant chemotherapy (NAC) prior to radical cystectomy is limited because of lack of tools that can guide for a better patient selection. We aim to explore if a combination of biomarkers is superior to a single marker. Pretreatment tumor specimens and clinical data from two randomized trials including 250 patients with T2-T4 urothelial bladder cancer, were used. The information on the expressions on tumor tissue of four biomarkers; CCTα, emmprin, survivin, and BCL-2, detected by immunohistochemistry in our previous studies, was used. Cox proportional hazard models, including treatment-by-biomarker interaction terms, were used to assess the predictive value of the biomarkers for efficacy of NAC on overall survival. CCTα provided predictive information about the efficacy of NAC (interaction P=0.009). None of the other biomarkers provided statistically significant information additional to CCTα. The adjusted hazard ratio for NAC treated versus no-NAC was 0.42 (95% CI: 0.27-0.64) for patients with negative CCTα expression, when adding information about emmprin it decreased to 0.33 (95% CI: 0.19-0.56) for patients with both negative CCTα and emmprin. This corresponds to a decrease in number needed to treat from 4 to 3 patients. The combination of CCTα with survivin or BCL-2 yielded similar results. In a group of patients with muscle invasive bladder cancer a combination of two biomarkers might improve the possibility to identify patients most likely to benefit from the use of NAC. Further studies designed to have sufficient power to detect an interaction effect are needed.

The "Burkitt-like" immunophenotype and genotype is rarely encountered in diffuse large B cell lymphoma and high-grade B cell lymphoma, NOS.

Burkitt lymphoma (BL) is a B cell lymphoma composed of monomorphic medium-sized blastic cells with basophilic cytoplasm and a high proliferation index. BL has a characteristic immunophenotype of CD10 and BCL6 positive and BCL2 negative and harbours MYC gene rearrangements (MYCR) in >90% of the cases. Owing to its highly aggressive nature, intensified chemotherapy regimens are usually administered, requiring an exact diagnosis. Since the diagnosis usually warrants an integration of morphologic, immunophenotypic and genetic findings and because there is a morphologic overlap with the new WHO category of high-grade B cell lymphoma, not otherwise specified (HGBL, NOS) and some cases of diffuse large B cell lymphoma (DLBCL), we wanted to test the distinctiveness of the CD10+, BCL6+, BCL2- and MYCR positive immunopheno-genotype in a large cohort of >1000 DLBCL and HGBL. Only 9/982 DLBCL classified by an expert panel of haematopathologists (0.9%) displayed a single MYCR and were CD10+, BCL6+ and BCL2-. In a similar fashion, only one out of 32 HGBL, NOS (3%) displayed the "Burkitt-like" genetic/immunophenotypic constitution. The samples of non-BL showing the BL-typic immunopheno-genotype, interestingly, harboured higher copy number variations (CNV) by OncoScan analysis (mean 7.3 CNVs/sample; range: 2-13 vs. 2.4; range 0-6) and were also distinct from pleomorphic BL cases regarding their mutational spectrum by NGS analysis. This implies that the characteristic immunophenotype of BL, in concert with a single MYCR, is uncommon in these aggressive lymphomas, and that this constellation favours BL.

Retrospective Analysis of Clinicopathological Characteristics of Lacrimal Gland Pleomorphic Adenoma and Mechanism of Tumorigenesis by the Imbalance Between Apoptosis and Proliferation.

BACKGROUND Lacrimal gland pleomorphic adenoma (LGPA) is the most common clinically benign epithelial tumor of the lacrimal gland and is predominantly comprised of epithelial cells and interstitial components. At present, the exact pathogenesis of LGPA remains unclear. Previous research has indicated that the occurrence of LGPA may be related to excessive cell proliferation. MATERIAL AND METHODS This study observed the clinicopathological characteristics of LGPA and investigated the tumorigenesis mechanism of cell over-proliferation caused by the imbalance between apoptosis and proliferation. A total of 27 cases were collected from the Department of Ophthalmology of the Affiliated Hospital of Chengde Medical University and the Third Medical Center of Chinese PLA General Hospital from April 2017 to November 2019. Hematoxylin-eosin (HE) staining and immunohistochemical staining were used to observe the pathological characteristics and analyze the expression of bcl-2 and bax in the lacrimal gland. RESULTS Compared with normal lacrimal gland tissues, LGPA tumor tissues had obvious changes in pathological morphology. The expression of bcl-2 in LGPA lesion tissues was dramatically higher (P<0.001), the expression of bax was not significantly different between groups (P=0.25), but the ratio of bcl-2/bax was significantly higher in tumor tissues (P=0.01). CONCLUSIONS We found that the lacrimal gland tumor tissues had obvious excessive proliferation in pathomorphology, which revealed the necessity of complete surgical removal of the capsule from the perspective of pathological morphology and provided a theoretical basis for the hypothesis that the imbalance between apoptosis and proliferation could lead to cell hyperproliferation.

Mitochondrial outer membrane permeabilization at the single molecule level.

Apoptotic cell death is essential for development, immune function or tissue homeostasis, and its mis-regulation is linked to various diseases. Mitochondrial outer membrane permeabilization (MOMP) is a central event in the intrinsic apoptotic pathway and essential to control the execution of cell death. Here we review current concepts in regulation of MOMP focusing on the interaction network of the Bcl-2 family proteins as well as further regulatory elements influencing MOMP. As MOMP is a complex spatially and temporally controlled process, we point out the importance of single-molecule techniques to unveil processes which would be masked by ensemble measurements. We report key single-molecule studies applied to decipher the composition, assembly mechanism and structure of protein complexes involved in MOMP regulation.

De novo design of modular and tunable protein biosensors.

Naturally occurring protein switches have been repurposed for the development of biosensors and reporters for cellular and clinical applications1. However, the number of such switches is limited, and reengineering them is challenging. Here we show that a general class of protein-based biosensors can be created by inverting the flow of information through de novo designed protein switches in which the binding of a peptide key triggers biological outputs of interest2. The designed sensors are modular molecular devices with a closed dark state and an open luminescent state; analyte binding drives the switch from the closed to the open state. Because the sensor is based on the thermodynamic coupling of analyte binding to sensor activation, only one target binding domain is required, which simplifies sensor design and allows direct readout in solution. We create biosensors that can sensitively detect the anti-apoptosis protein BCL-2, the IgG1 Fc domain, the HER2 receptor, and Botulinum neurotoxin B, as well as biosensors for cardiac troponin I and an anti-hepatitis B virus antibody with the high sensitivity required to detect these molecules clinically. Given the need for diagnostic tools to track the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)3, we used the approach to design sensors for the SARS-CoV-2 spike protein and antibodies against the membrane and nucleocapsid proteins. The former, which incorporates a de novo designed spike receptor binding domain (RBD) binder4, has a limit of detection of 15 pM and a luminescence signal 50-fold higher than the background level. The modularity and sensitivity of the platform should enable the rapid construction of sensors for a wide range of analytes, and highlights the power of de novo protein design to create multi-state protein systems with new and useful functions.