Thrombopoietin receptor agonists for refractory thrombocytopenia in patients after autologous hematopoietic stem cell transplantation.

OBJECTIVE: Autologous hematopoietic stem cell (ASC) transplantation (ASCT) is an effective treatment method for patients with hematological disorders and malignant diseases. The patient's ASCs are harvested prior to radiotherapy/chemotherapy, cryopreserved and then transfused back after the high-dose radiotherapy/chemotherapy conditioning treatment. Since some patients develop thrombocytopenia after receiving ASCT, it is difficult for them to bear simultaneously the management of their original disease and thrombocytopenia. The present study aimed to evaluate the efficacy and safety of thrombocytopenia therapy with thrombopoietin receptor agonists (TPORAs) after ASCT. METHODS: We retrospectively analyzed the clinical safety and efficacy of TPORA treatment for the enrolled 20 patients who developed thrombocytopenia after ASCT. The measured parameters were prolonged isolated thrombocytopenia (PIT), secondary failure of platelet recovery (SFPR) and other calculated response index. Patients with platelet count (PC) ≤ 50×109/L were treated with TPORA, namely with either eltrombopag (Elt), hetrombopag (Het), or avatrobopag (Ava). RESULTS: The group of 20 patients, who received TPORA administration for their thrombocytopenia after ASCT, had a median age of 50 years (ranging between 17 and 60 years). The median administration time of TPORA application was 48 days (ranging from 7 to 451 days); an overall response rate (ORR) was 85% with no response in 15% of patients, while with complete response (CR) in 70% of patients and partial response (PR) in 15% of patients. The median platelet count was 19 × 109/L before TPORA treatment and increased to 87×109)/L after the treatment. The TPORA treatment was safe as only 4 patients (20%) displayed a mild transaminase elevation. No other reported side effects occurred, such as thrombosis, joint pain, diarrhea, and myelofibrosis. It was demonstrated that the short response time to TPORA treatment correlated to the fast platelet recovery, when the number of megakaryocytes in the bone marrow smear exceeded 35/4.5 cm2 under a low magnification of 100 times (p = 0.015). CONCLUSION: TPORA therapy for thrombocytopenia occurring after the radiotherapy/ chemotherapy-conditioned ASCT was well tolerated and effective for platelets recovery.

HSP90/CDC37 inactivation promotes degradation of LKB1 protein to suppress AMPK signaling in bronchial epithelial cells exposed to sulfur mustard analog, 2-chloroethyl ethyl sulfide.

To investigate the role of the liver kinase (LK) B1 protein, an activator of AMP-activated protein kinase (AMPK), in AMPK signaling suppression when exposed to vesicant, a kind of chemical warfare agent. Cultured human bronchial epithelial cells were inflicted with sulfur mustard (SM) analog, 2-chloroethyl ethyl sulfide (CEES) of 0.2-1.0 mM concentration, and cell proliferation, apoptosis, autophagy, and cellular ATP level were analyzed up to 24 h after the exposure. Focusing on LKB1, heat shock protein (HSP) 90, and cell division cycle (CDC) 37 proteins, the protein expression, phosphorylation, and interaction were examined with western blot, immunofluorescence staining, and/or immunoprecipitation. AMPK signaling was found to be inhibited 24 h after being exposed to either sub-cytotoxic (0.5 mM) or cytotoxic (1.0 mM) concentration of CEES based on MTS assay. Consistently, the degradation of the LKB1 protein and its less interaction with the HSP90/CDC37 complex was confirmed. It was found that 1.0, not 0.5 mM CEES also decreased the CDC37 protein, proteasome activity, and cellular ATP content that modulates HSP90 protein conformation. Inhibiting proteasome activity could alternatively activate autophagy. Finally, either 0.5 or 1.0 mM CEES activated HSP70 and autophagy, and the application of an HSP70 inhibitor blocked autophagy and autophagic degradation of the LKB1 protein. In conclusion, we reported here that AMPK signaling inactivation by CEES was a result of LKB1 protein loss via less protein complex formation and enhanced degradation.

Azacitidine and lenalidomide combination: a novel relapse prophylaxis regimen after allogeneic hematopoietic stem-cell transplantation in patients with acute myeloid leukemia.

Introduction: While allogeneic hematopoietic stem cell transplantation (allo-HSCT) can be a curative regimen for acute myeloid leukemia (AML), relapse of AML remains a serious risk post-transplantation. Once relapsed, salvage options are limited and management of AML is difficult. Here we designed a prospective study to examine the efficacy and tolerability of maintenance therapy with azacytidine (AZA) plus low-dose lenalidomide (LEN) to prevent relapse after allo-HSCT for AML patients (ChiCTR2200061803). Methods: AML patients post-allo-HSCT were treated with AZA (75 mg/m2 for 7 days), followed by LEN (5 mg/m2, day 10-28), and a 4-week resting interval, which was defined as one treatment cycle. A total of 8 cycles was recommended. Results: 37 patients were enrolled, 25 patients received at least 5 cycles, and 16 patients finished all 8 cycles. With a median follow-up time of 608 (43-1440) days, the estimated 1-year disease free survival (DFS) was 82%, cumulative incidence of relapse (CIR) was 18%, and overall survival (OS) was 100%. Three patients (8%) had grade 1-2 neutropenia without fever; one patient developed grade 3-4 thrombocytopenia and minor subdural hematoma; 4/37 patients (11%) developed chronic GVHD with a score of 1-2, without requiring systemic treatment; No patient developed acute GVHD. After AZA/LEN prophylaxis, increasing numbers of CD56+NK and CD8+ T, and decreasing of CD19+ B cells were observed. Discussion: Azacitidine combined with low-dose lenalidomide was observed to be an effective relapse prophylaxis option after allo-HSCT in AML patients, and can be administered safely without significantly increasing the risk of GVHD, infection and other AEs. Clinical Trial Registration: www.chictr.org, identifier ChiCTR2200061803.

Sulfur mustard analog 2-chloroethyl ethyl sulfide increases triglycerides by activating DGAT1-dependent biogenesis and inhibiting PGC1ɑ-dependent fat catabolism in immortalized human bronchial epithelial cells.

Using sulfur mustard analog 2-chloroethyl ethyl sulfide (CEES), we established an in vitro model by poisoning cultured immortalized human bronchial epithelial cells. Nile Red staining revealed lipids accumulated 24 h after a toxic dose of CEES (0.9 mM). Lipidomics analysis showed most of the increased lipids were triglycerides (TGs), and the increase in TGs was further confirmed using a Triglyceride-Glo™ Assay kit. Protein and mRNA levels of DGAT1, an important TG biogenesis enzyme, were increased following 0.4 mM CEES exposure. Under higher dose CEES (0.9 mM) exposure, protein and mRNA levels of PPARγ coactivator-1ɑ (PGC-1ɑ), a well-known transcription factor that regulates fatty acid oxidation, were decreased. Finally, application with DGAT1 inhibitor A 922500 or PGC1ɑ agonist ZLN005 was able to block the CEES-induced TGs increase. Overall, our dissection of CEES-induced TGs accumulation provides new insight into energy metabolism dysfunction upon vesicant exposure.HIGHLIGHTSIn CEES (0.9 mM)-injured cells:Triglycerides (TGs) were abundant in the accumulated lipids.Expression of DGAT1, not DGAT2, was increased.Expression of PGC1ɑ, not PGC1ß, was reduced.DGAT1 inhibitor or PGC1ɑ agonist blocked the CEES-mediated increase in TGs.

Pretransplantation minimal residual disease monitoring by multiparameter flow cytometry predicts outcomes of AML patients receiving allogeneic hematopoietic stem cell transplantation.

BACKGROUND AND PURPOSE: Is minimal residual disease (MRD) monitoring by multiparameter flow cytometry (MFC) prognostic for acute myeloid leukemia (AML) patients before allogeneic hemopoietic stem cell transplantation (allo-HSCT)? And if so, what level of MRD eradication can be used to help guide the timing of HSCT? Can haplo-HSCT improve the prognosis of AML patients with MRD positive? To figure out these questions, we initiated this retrospective study. METHODS: 96 AML patients were included retrospectively and divided into 5 groups, according to pre-transplantation MRD levels (from 5 × 10-2 to <1 × 10-4), to analyze the overall survival (OS), disease-free survival (DFS) and cumulative incidence of relapse (CIR). Secondly, we compared the prognosis of MRD-negative (MRDneg) and MRD-positive (MRDpos) AML patients (cutoff value = 1 × 10-3) who underwent allo-HSCT, and further analyzed the prognosis of MRDpos patients after received different transplantation modalities. RESULTS: It is found that the 2-year OS and DFS of MRD negative group were better than the MRD positive group, and that the deeper the eradication of MRD before transplantation, the better the prognosis of patients. The CIR in patients received HLA-identical transplantation, was higher in the MRDpos than in the MRDneg. Haploid transplantation reduced the CIR disparity between MRDpos and MRDneg group. Subsequently, in AML patients who remain MRD positive before HSCT, we show that haplo-HSCT offered a better prognosis than HLA-identical transplantation (MSDT and MUDT). CONCLUSION: It is suggested that achieving MFC-MRD <10-3 (10-4 or even better) before allo-HSCT could reduce the relapse of AML and improve OS and DFS significantly, while haplo-HSCT may be preferred for patients not achieving MRD negativity.

Vitamin D3 protects against nitrogen mustard-induced apoptosis of the bronchial epithelial cells via activating the VDR/Nrf2/Sirt3 pathway.

Respiratory system injury is the main cause of mortality for nitrogen mustard (NM)-induced damage. Previous studies indicate that reactive oxygen species (ROS) participates in NM-mediated respiratory injuries, but the detailed mechanism is not quite clear. Human bronchial epithelial cell lines 16HBE and BEAS-2B were treated with HN2, a type of NM. In detail, it was shown that HN2 treatment induced impaired cell viability, excessive mitochondrial ROS production and enhanced cellular apoptosis in bronchial epithelial cells. Moreover, impaired Sirt3/SOD2 axis was observed upon HN2 treatment, with decreased Sirt3 and increased acetylated SOD2 expression levels. Sirt3 overexpression partially ameliorated HN2-induced cell injury. Meanwhile, vitamin D3 treatment partially attenuated HN2-induced apoptosis and improved the mitochondrial functions upon HN2 intervention. In addition, HN2 exposure decreased VDR expression, thus inhibiting the Nrf2 phosphorylation and Sirt3 activation. Inhibition of Nrf2 or Sirt3 could decrease the protective effects of vitamin D3 and enhance mitochondrial ROS production via modulating mitochondrial redox balance. In conclusion, impaired VDR/Nrf2/Sirt3 axis contributed to NM-induced apoptosis, while vitamin D3 supplementation provides protective effects via the activation of VDR and the improvement of mitochondrial functions. This study provides novel mechanism and strategy for NM exposure-induced pulmonary injuries.

Case Report: PD-1 Blockade Combined Autologous Hematopoietic Stem Cell Transplantation With Modified BEAM Regimen Containing High-Dose Cytarabine to Treat R/R Hodgkin's Lymphoma.

The emergence of new drugs has provided additional options in the treatment of relapsed and refractory (R/R) Hodgkin's lymphoma (HL). However, the use of autologous stem cell transplantation (ASCT) has not been completely replaced in this setting. The use of anti-programmed death-1 (PD-1) antibody bridging to ASCT and as maintenance after transplantation is a novel approach in HL treatment. In this case, we report that PD-1 monoclonal antibody (mAb) plus ASCT with modified BEAM regimen (carmustine + etoposide + cytarabine + melphalan) containing high-dose cytarabine to treat R/R HL may represent a promising regimen in this difficult-to-treat setting.

Critical role of cysteine-266 of SIE3 in regulating the ubiquitination and degradation of SIP1 transcription factor in Lotus japonicus.

MAIN CONCLUSION: A conserved cysteine residue (C266)-mediated homo-dimerization of SIE3 is required for the ubiquitination and degradation of SIP1 transcription factor in Lotus japonicas CTLH/CRA/RING-containing proteins have been shown to possess E3-ligase activities and are crucial for the regulation of numerous cellular signaling pathways. In our previous studies, SIE3 (SymRK-Interacting E3 ubiquitin ligase), a CTLH/CRA/RING-containing protein from Lotus japonicus, has been shown to associate with both Symbiosis Receptor Kinase (SymRK) and SIP1 (SymRK interacting protein 1) transcription factor, and ubiquitinate SymRK (Yuan et al. Plant Physiol 160 (1):106-117, 2012; Feng et al. Front Plant Sci 11: 795, 2020). Besides, we previously also demonstrated that the residue, cysteine-266 in the CRA (CT11-RanBPM) domain is required for homodimerization of SIE3 and cysteine-266 residue-mediated homodimerization is important for the symbiosic function of SIE3 (Feng et al. 2020). In this report, SIE3 was shown to induce the ubiquitination and degradation of SIP1. The cysteine-266 residue is essential for the E3-ligase activity and is highly conserved in the SIE3-like proteins. Our works refined the working model that homodimerization of SIE3 is required for ubiquitin-related degradation of SIP1 and found a conserved cysteine residue plays a key role in the activity of a plant dimeric E3 ligase.

PGC-1ɑ Mediated-EXOG, a Specific Repair Enzyme for Mitochondrial DNA, Plays an Essential Role in the Rotenone-Induced Neurotoxicity of PC12 Cells.

Mitochondria harbor small circular genomes (mtDNA) that encode 13 oxidative phosphorylation (OXPHOS) proteins, and types of damage to mtDNA may contribute to neuronal damage. Recent studies suggested that regulation of mtDNA repair proteins may be a potential strategy for treating neuronal damage. The mtDNA repair system contains its own repair enzymes and is independent from the nuclear DNA repair system. Endo/exonuclease G-like(EXOG) is a mitochondria-specific 5-exo/endonuclease required for repairing endogenous single-strand breaks (SSBs) in mtDNA. However, whether EXOG plays a key role in neuronal damage induced by rotenone remains unknown. Thus, in this study, we aimed to investigate the effect of EXOG on mtDNA repair and mitochondrial functional maintenance in rotenone-induced neurotoxicity. Our results indicated that rotenone influenced the expression and location of EXOG in PC12 cells. Meanwhile, after rotenone exposure, the expression was reduced for proteins responsible for mtDNA repair, including DNA polymerase γ (POLG), high-temperature requirement protease A2 (HtrA2), and the heat-shock factor 1-single-stranded DNA-binding protein 1 (HSF1-SSBP1) complex. Further analysis demonstrated that EXOG knockdown led to reduced mtDNA copy number and mtDNA transcript level and increased mtDNA deletion, which further aggravated the mtDNA damage and mitochondrial dysfunction under rotenone stress. In turn, EXOG overexpression protected PC12 cells from mtDNA damage and mitochondrial dysfunction induced by rotenone. As a result, EXOG knockdown reduced cell viability and tyrosine hydroxylase expression, while EXOG overexpression alleviated rotenone's effect on cell viability and tyrosine hydroxylase expression in PC12 cells. Further, we observed that EXOG influenced mtDNA repair by regulating protein expression of the HSF1-SSBP1 complex and POLG. Furthermore, our study showed that PGC-1α upregulation with ZLN005 led to increased protein levels of EXOG, POLG, HSF1, and SSBP1, all of which contribute to mtDNA homeostasis. Therefore, PGC-1α may be involved in mtDNA repair through interacting with multiple mtDNA repair proteins, especially with the help of EXOG. In summary, EXOG regulation by PGC-1α plays an essential role in rotenone-induced neurotoxicity in PC12 cells. EXOG represents a protective effect strategy in PC12 cells exposed to rotenone.

The Role of mTOR Inhibitors in Hematologic Disease: From Bench to Bedside.

The mTOR pathway plays a central role in many cellular processes, such as cellular growth, protein synthesis, glucose, and lipid metabolism. Aberrant regulation of mTOR is a hallmark of many cancers, including hematological malignancies. mTOR inhibitors, such as Rapamycin and Rapamycin analogs (Rapalogs), have become a promising class of agents to treat malignant blood diseases-either alone or in combination with other treatment regimens. This review highlights experimental evidence underlying the molecular mechanisms of mTOR inhibitors and summarizes their evolving role in the treatment of hematologic disease, including leukemia, lymphoma, myeloma, immune hemocytopenia, and graft-versus-host disease (GVHD). Based on data presented in this review, we believe that mTOR inhibitors are becoming a trusted therapeutic in the clinical hematologist's toolbelt and should be considered more routinely in combination therapy for the management of hematologic disease.

Nitrogen mustard prevents transport of Fra-1 into the nucleus to promote c-Fos- and FosB-dependent IL-8 induction in injured mouse epidermis.

Transcription factor activator protein (AP)-1 can be activated in nitrogen-mustard-injured mouse skin, and is thought to participate in the inflammatory response. AP-1 consists of homo- or heterodimers of Fos [c-Fos, Fos-B, fos-related antigen (Fra)-1 and Fra-2] and Jun (c-Jun, JunB and JunD) family members, and information about their expression, location and function are still unclear. In nitrogen-mustard-exposed mouse skin, we found p-ERK activation increased Fra-1 and FosB. Unlike the nucleus location of c-Fos and FosB, Fra-1 and Fra-2 were mainly expressed in the cytoplasm. In nitrogen-mustard-exposed cultured immortalized human keratinocytes (HaCaT cells), Fra-1 in the nucleus functioned as an inhibitor of inflammatory cytokine interleukin (IL)-8. Co-immunoprecipitation showed that Fra-1 formed dimers with IL-8 transcription factors c-Jun, JunB and JunD. Fra-1 depletion increased c-Fos and FosB in the nucleus, accompanied by increased heterodimers of c-Fos/c-Jun, c-Fos/JunB, c-Fos/JunD, and FosB/JunB. In conclusion, Fra-1 trapped in the cytoplasm after nitrogen mustard exposure might be a driving force for IL-8 over-expression in injured skin.

TET2 Function in Hematopoietic Malignancies, Immune Regulation, and DNA Repair.

Over the last decade, investigation of Ten-Eleven Translocation 2 (TET2) gene function and TET2 mutation have become of increasing interest in the field of hematology. This heightened interest was sparked by the seminal discoveries that (1) TET2 mutation is associated with development of hematological malignancies and that (2) the TET family of proteins is critical in promoting DNA demethylation and immune homeostasis. Since then, additional studies have begun to unravel the question "Does TET2 have additional biological functions in the regulation of hematopoiesis?" Here, we present a mini-review focused on the current understanding of TET2 in hematopoiesis, hematological malignancies, and immune regulation. Importantly, we highlight the critical function that TET2 facilitates in maintaining the stability of the genome. Based on our review of the literature, we provide a new hypothesis that loss of TET2 may lead to dysregulation of the DNA repair response, augment genome instability, and subsequently sensitize myeloid leukemia cells to PARP inhibitor treatment.

Mitochondrial ATP-sensitive potassium channel regulates mitochondrial dynamics to participate in neurodegeneration of Parkinson's disease.

Parkinson's disease (PD) is the second most common age-related neurodegenerative disease. Mitochondrial dysfunction has been the focus of the pathogenesis of PD. The mitochondrial ATP-sensitive potassium channel (mitoKATP) plays a significant role in mitochondrial physiology and has been extensively shown to protect against ischemic and brain reperfusion injury. However, there have long been controversies regarding its role in Parkinson's disease. We investigated the role of mitoKATP channels in rotenone-induced PD model in vivo and vitro and the interactions of mitoKATP channels, mitochondrial dynamics and PD. The results indicated that the use of diazoxide to activate mitoKATP channels resulted in the aggravation of rotenone-induced dopamine neurodegeneration in PC12 cells and SD rats. In contrast, the use of 5-hydroxydecanoate (5-HD) to inhibit mitoKATP channels improved rotenone-induced dopamine neurodegeneration, which was not consistent with mitoKATP channels in ischemic and brain reperfusion injury. Further analysis determined that the mitoKATP channel was involved in PD mainly via the regulation of mitochondrial biogenesis and fission/fusion. And the pore subunits of Kir6.1, the major component of mitoKATP channels, was the key contributor in its interaction with mitochondrial dynamics in rotenone-induced dopamine neurodegeneration. Therefore, it can be concluded that mitoKATP channels regulate mitochondrial dynamics to participate in rotenone-induced PD mainly attributes to the pore subunits of Kir6.1. And additionally, though mitoKATP channels may represent a direction of one potential target for neuroprotection, it should be noted that the effects are different in the activation or inhibition of mitoKATP channels in different models.

Formation and degradation of nitrogen mustard-induced MGMT-DNA crosslinking in 16HBE cells.

N-methyl-2,2-di(chloroethyl)amine (HN2) is a kind of bifunctional alkyltating agent, which can react with nucleophilic groups in DNA and/or protein to form HN2-bridged crosslinking of target molecules, such as DNA-protein crosslinkings (DPC). O6-methylguanine-DNA methyltransferase (MGMT) is a DNA damage repair enzyme which solely repairs alkyl adduct on DNA directly. However, MGMT was detected to act as a protein cross-linked with DNA via alkylation in presence of HN2, and unexpectedly turned into a DNA damage enhancer in the form of MGMT-DNA cross-link (mDPC). Present study aimed to explore the possible ways to lessen the incorporation of MGMT into DPC as well as to save it for DNA repair. To find out the influencing factors of mDPC formation and cleavage, human bronchial epithelial cell line 16HBE was exposed to HN2 and the factors related with MGMT expression and degradation were investigated. When c-Myc, a negative transcriptional factor of MGMT was inhibited by 10058-F4, MGMT expression and mDPC formation were increased, and more γ-H2AX was also detected. Sustained treatment with O6BG, a specific exogenous substrate and depleter of MGMT, could reduce the level of MGMT and mDPC formation. In contrast, a transient 1h pre-treatment of O6GB before HN2 exposure would cause a high MGMT and mDPC level. MGMT was increasingly ubiquitinated after HN2 exposure in a time-dependent manner. At the same time, MGMT was also SUMOylated with a downward time-dependent manner compared to its ubiquitination. Inhibitors of E1, E2 or E3 ligases of ubiqutination all led to the accumulation of mDPC and total-DPC (tDPC) with the difference as that mDPC was sensitive to E1 inhibitor while tDPC more sensitive to E2 and E3 inhibitor. Our results demonstrated the control of mDPC level could be realized through transcription inhibitory effect of c-Myc, O6GB application, and the acceleration of mDPC ubiquitination and subsequent degradation.

Cytoplasmic expression of C-MYC protein is associated with risk stratification of mantle cell lymphoma.

AIM: To investigate the association of C-MYC protein expression and risk stratification in mantle cell lymphoma (MCL), and to evaluate the utility of C-MYC protein as a prognostic biomarker in clinical practice. METHODS: We conducted immunohistochemical staining of C-MYC, Programmed cell death ligand 1 (PD-L1), CD8, Ki-67, p53 and SRY (sex determining region Y) -11 (SOX11) to investigate their expression in 64 patients with MCL. The staining results and other clinical data were evaluated for their roles in risk stratification of MCL cases using ANOVA, Chi-square, and Spearman's Rank correlation coefficient analysis. RESULTS: Immunohistochemical staining in our study indicated that SOX11, Ki-67 and p53 presented nuclear positivity of tumor cells, CD8 showed membrane positivity in infiltrating T lymphocytes while PD-L1 showed membrane and cytoplasmic positivity mainly in macrophage cells and little in tumor cells. We observed positive staining of C-MYC either in the nucleus or cytoplasm or in both subcellular locations. There were significant differences in cytoplasmic C-MYC expression, Ki-67 proliferative index of tumor cells, and CD8 positive tumor infiltrating lymphocytes (CD8+TIL) among three risk groups (P = 0.000, P = 0.037 and P=0.020, respectively). However, no significant differences existed in the expression of nuclear C-MYC, SOX11, p53, and PD-L1 in MCL patients with low-, intermediate-, and high risks. In addition, patient age and serum LDH level were also significantly different among 3 groups of patients (P = 0.006 and P = 0.000, respectively). Spearman's rank correlation coefficient analysis indicated that cytoplasmic C-MYC expression, Ki-67 index, age, WBC, as well as LDH level had significantly positive correlations with risk stratification (P = 0.000, 0.015, 0.000, 0.029 and 0.000, respectively), while CD8+TIL in tumor microenvironment negatively correlated with risk stratification of patients (P = 0.006). Patients with increased positive cytoplasmic expression of C-MYC protein and decreased CD8+TIL appeared to be associated with a poor response to chemotherapy, but the correlation was not statistically significant. CONCLUSION: Our study suggested that assessment of cytoplasmic C-MYC overexpression and cytotoxic T lymphocytes (CTLs) by immunohistochemical staining might be helpful for MCL risk stratification and outcome prediction. However, large cohort studies of MCL patients with complete follow up are needed to validate our speculation.

The Interaction of Mitochondrial Biogenesis and Fission/Fusion Mediated by PGC-1α Regulates Rotenone-Induced Dopaminergic Neurotoxicity.

Parkinson's disease is a common neurodegenerative disease in the elderly, and mitochondrial defects underlie the pathogenesis of PD. Impairment of mitochondrial homeostasis results in reactive oxygen species formation, which in turn can potentiate the accumulation of dysfunctional mitochondria, forming a vicious cycle in the neuron. Mitochondrial fission/fusion and biogenesis play important roles in maintaining mitochondrial homeostasis. It has been reported that PGC-1α is a powerful transcription factor that is widely involved in the regulation of mitochondrial biogenesis, oxidative stress, and other processes. Therefore, we explored mitochondrial biogenesis, mitochondrial fission/fusion, and especially PGC-1α as the key point in the signaling mechanism of their interaction in rotenone-induced dopamine neurotoxicity. The results showed that mitochondrial number and mass were reduced significantly, accompanied by alterations in proteins known to regulate mitochondrial fission/fusion (MFN2, OPA1, Drp1, and Fis1) and mitochondrial biogenesis (PGC-1α and mtTFA). Further experiments proved that inhibition of mitochondrial fission or promotion of mitochondrial fusion has protective effects in rotenone-induced neurotoxicity and also promotes mitochondrial biogenesis. By establishing cell models of PGC-1α overexpression and reduced expression, we found that PGC-1α can regulate MFN2 and Drp1 protein expression and phosphorylation to influence mitochondrial fission/fusion. In summary, it can be concluded that PGC-1α-mediated cross talk between mitochondrial biogenesis and fission/fusion contributes to rotenone-induced dopaminergic neurodegeneration.

Massive Cerebrospinal Fluid Replacement Reduces Delayed Cerebral Vasospasm After Embolization of Aneurysmal Subarachnoid Hemorrhage.

BACKGROUND Delayed cerebral vasospasm (DCVS) following aneurismal subarachnoid hemorrhage (SAH) is a leading cause of poor prognosis and death in SAH patients. Effective management to reduce DCVS is needed. A prospective controlled trial was conducted to determine if massive cerebrospinal fluid (CSF) replacement (CR) could reduce DCVS occurrence and improve the clinical outcome after aneurysmal SAH treated with endovascular coiling. MATERIAL AND METHODS Patients treated with endovascular coiling after aneurysmal SAH were randomly divided into a control group receiving regular therapy alone (C group, n=42) and a CSF replacement group receiving an additional massive CSF replacement with saline (CR group, n=45). CSF examination, head CT, DCVS occurrence, cerebral infarction incidence, Glasgow Outcome Scale prognostic score, and 1-month mortality were recorded. RESULTS The occurrence of DCVS was 30.9% in the C group and 4.4% in the CR group (P<0.005). The cerebral infarction incidences in the C and CR groups were 19.0% and 2.2% (P<0.05), respectively, 1 month after the treatments. Mortality was not significantly different between the 2 groups during the follow-up period. CONCLUSIONS Massive CR after embolization surgery for aneurysmal SAH can significantly reduce DCVS occurrence and effectively improve the outcomes.

Bifunctional alkylating agent-mediated MGMT-DNA cross-linking and its proteolytic cleavage in 16HBE cells.

Nitrogen mustard (NM), a bifunctional alkylating agent (BAA), contains two alkyl arms and can act as a cross-linking bridge between DNA and protein to form a DNA-protein cross-link (DPC). O(6)-methylguanine-DNA methyltransferase (MGMT), a DNA repair enzyme for alkyl adducts removal, is found to enhance cell sensitivity to BAAs and to promote damage, possibly due to its stable covalent cross-linking with DNA mediated by BAAs. To investigate MGMT-DNA cross-link (mDPC) formation and its possible dual roles in NM exposure, human bronchial epithelial cell line 16HBE was subjected to different concentrations of HN2, a kind of NM, and we found mDPC was induced by HN2 in a concentration-dependent manner, but the mRNA and total protein of MGMT were suppressed. As early as 1h after HN2 treatment, high mDPC was achieved and the level maintained for up to 24h. Quick total DPC (tDPC) and γ-H2AX accumulation were observed. To evaluate the effect of newly predicted protease DVC1 on DPC cleavage, we applied siRNA of MGMT and DVC1, MG132 (proteasome inhibitor), and NMS-873 (p97 inhibitor) and found that proteolysis plays a role. DVC1 was proven to be more important in the cleavage of mDPC than tDPC in a p97-dependent manner. HN2 exposure induced DVC1 upregulation, which was at least partially contributed to MGMT cleavage by proteolysis because HN2-induced mDPC level and DNA damage was closely related with DVC1 expression. Homologous recombination (HR) was also activated. Our findings demonstrated that MGMT might turn into a DNA damage promoter by forming DPC when exposed to HN2. Proteolysis, especially DVC1, plays a crucial role in mDPC repair.

Resveratrol Regulates Mitochondrial Biogenesis and Fission/Fusion to Attenuate Rotenone-Induced Neurotoxicity.

It has been confirmed that mitochondrial impairment may underlie both sporadic and familial Parkinson's disease (PD). Mitochondrial fission/fusion and biogenesis are key processes in regulating mitochondrial homeostasis. Therefore, we explored whether the protective effect of resveratrol in rotenone-induced neurotoxicity was associated with mitochondrial fission/fusion and biogenesis. The results showed that resveratrol could not only promote mitochondrial mass and DNA copy number but also improve mitochondrial homeostasis and neuron function in rats and PC12 cells damaged by rotenone. We also observed effects with alterations in proteins known to regulate mitochondrial fission/fusion and biogenesis in rotenone-induced neurotoxicity. Therefore, our findings suggest that resveratrol may prevent rotenone-induced neurotoxicity through regulating mitochondrial fission/fusion and biogenesis.

Cell Density-Dependent Upregulation of PDCD4 in Keratinocytes and Its Implications for Epidermal Homeostasis and Repair.

Programmed cell death 4 (PDCD4) is one multi-functional tumor suppressor inhibiting neoplastic transformation and tumor invasion. The role of PDCD4 in tumorigenesis has attracted more attention and has been systematically elucidated in cutaneous tumors. However, the normal biological function of PDCD4 in skin is still unclear. In this study, for the first time, we find that tumor suppressor PDCD4 is uniquely induced in a cell density-dependent manner in keratinocytes. To determine the potential role of PDCD4 in keratinocyte cell biology, we show that knockdown of PDCD4 by siRNAs can promote cell proliferation in lower cell density and partially impair contact inhibition in confluent HaCaT cells, indicating that PDCD4 serves as an important regulator of keratinocytes proliferation and contact inhibition in vitro. Further, knockdown of PDCD4 can induce upregulation of cyclin D1, one key regulator of the cell cycle. Furthermore, the expression patterns of PDCD4 in normal skin, different hair cycles and the process of wound healing are described in detail in vivo, which suggest a steady-state regulatory role of PDCD4 in epidermal homeostasis and wound healing. These findings provide a novel molecular mechanism for keratinocytes' biology and indicate that PDCD4 plays a role in epidermal homeostasis.