Browsing by Subject "apoptosis"

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  • Buettner, Ralf; Le Xuan Truong Nguyen; Kumar, Bijender; Morales, Corey; Liu, Chao; Chen, Lisa S.; Pemovska, Tea; Synold, Timothy W.; Palmer, Joycelynne; Thompson, Ryan; Li, Ling; Dinh Hoa Hoang; Zhang, Bin; Ghoda, Lucy; Kowolik, Claudia; Kontro, Mika; Leitch, Calum; Wennerberg, Krister; Yu, Xiaochun; Chen, Ching-Cheng; Horne, David; Gandhi, Varsha; Pullarkat, Vinod; Marcucci, Guido; Rosen, Steven T. (2019)
    Nucleoside analogs represent the backbone of several distinct chemotherapy regimens for acute myeloid leukemia (AML) and combination with tyrosine kinase inhibitors has improved survival of AML patients, including those harboring the poor-risk FLT3-ITD mutation. Although these compounds are effective in killing proliferating blasts, they lack activity against quiescent leukemia stem cells (LSCs), which contributes to initial treatment refractoriness or subsequent disease relapse. The reagent 8-chloro-adenosine (8-Cl-Ado) is a ribose-containing, RNA-directed nucleoside analog that is incorporated into newly transcribed RNA rather than in DNA, causing inhibition of RNA transcription. In this report, we demonstrate antileukemic activities of 8-Cl-Ado in vitro and in vivo and provide mechanistic insight into the mode of action of 8-Cl-Ado in AML. 8-Cl-Ado markedly induced apoptosis in LSC, with negligible effects on normal stem cells. 8-Cl-Ado was particularly effective against AML cell lines and primary AML blast cells harboring the FLT3-ITD mutation. FLT3-ITD is associated with high expression of miR-155. Furthermore, we demonstrate that 8-Cl-Ado inhibits miR-155 expression levels accompanied by induction of DNA-damage and suppression of cell proliferation, through regulation of miR-155/ErbB3 binding protein 1(Ebp1)/p53/PCNA signaling. Finally, we determined that combined treatment of NSG mice engrafted with FLT3-ITD (+) MV4-11 AML cells with 8-Cl-Ado and the FLT3 inhibitor AC220 (quizartinib) synergistically enhanced survival, compared with that of mice treated with the individual drugs, suggesting a potentially effective approach for FLT3-ITD AML patients.
  • Kakkola, L.; Denisova, O. V.; Tynell, J.; Viiliainen, J.; Ysenbaert, T.; Matos, R. C.; Nagaraj, A.; Öhman, Tiina; Kuivanen, S.; Paavilainen, H.; Feng, L.; Yadav, B.; Julkunen, I.; Vapalahti, O.; Hukkanen, V.; Stenman, J.; Aittokallio, T.; Verschuren, E. W.; Ojala, P. M.; Nyman, T.; Saelens, X.; Dzeyk, K.; Kainov, D. E. (2013)
  • Bulanova, Daria; Ianevski, Aleksandr; Bugai, Andrii; Akimov, Yevhen; Kuivanen, Suvi; Paavilainen, Henrik; Kakkola, Laura; Nandania, Jatin; Turunen, Laura; Ohman, Tiina; Ala-Hongisto, Hanna; Pesonen, Hanna M.; Kuisma, Marika S.; Honkimaa, Anni; Walton, Emma L.; Oksenych, Valentyn; Lorey, Martina B.; Guschin, Dmitry; Shim, Jungmin; Kim, Jinhee; Than, Thoa T.; Chang, So Young; Hukkanen, Veijo; Kulesskiy, Evgeny; Marjomaki, Varpu S.; Julkunen, Ilkka; Nyman, Tuula A.; Matikainen, Sampsa; Saarela, Jani S.; Sane, Famara; Hober, Didier; Gabriel, Guelsah; De Brabander, Jef K.; Martikainen, Miika; Windisch, Marc P.; Min, Ji-Young; Bruzzone, Roberto; Aittokallio, Tero; Vaha-Koskela, Markus; Vapalahti, Olli; Pulk, Arto; Velagapudi, Vidya; Kainov, Denis E. (2017)
    Viral diseases remain serious threats to public health because of the shortage of effective means of control. To combat the surge of viral diseases, new treatments are urgently needed. Here we show that small-molecules, which inhibit cellular anti-apoptotic Bcl-2 proteins (Bcl-2i), induced the premature death of cells infected with different RNA or DNA viruses, whereas, at the same concentrations, no toxicity was observed in mock-infected cells. Moreover, these compounds limited viral replication and spread. Surprisingly, Bcl-2i also induced the premature apoptosis of cells transfected with viral RNA or plasmid DNA but not of mock-transfected cells. These results suggest that Bcl-2i sensitizes cells containing foreign RNA or DNA to apoptosis. A comparison of the toxicity, antiviral activity, and side effects of six Bcl-2i allowed us to select A-1155463 as an antiviral lead candidate. Thus, our results pave the way for the further development of Bcl-2i for the prevention and treatment of viral diseases.
  • Lantto, Tiina A.; Laakso, Into; Dorman, H. J. Damien; Mauriala, Timo; Hiltunen, Raimo; Köks, Sulev; Raasmaja, Atso (2016)
    Plant phenolics have shown to activate apoptotic cell death in different tumourigenic cell lines. In this study, we evaluated the effects of juniper berry extract (Juniperus communis L.) on p53 protein, gene expression and DNA fragmentation in human neuroblastoma SH-SY5Y cells. In addition, we analyzed the phenolic composition of the extract. We found that juniper berry extract activated cellular relocalization of p53 and DNA fragmentation-dependent cell death. Differentially expressed genes between treated and non-treated cells were evaluated with the cDNA-RDA (representational difference analysis) method at the early time point of apoptotic process when p53 started to be activated and no caspase activity was detected. Twenty one overexpressed genes related to cellular stress, protein synthesis, cell survival and death were detected. Interestingly, they included endoplasmic reticulum (ER) stress inducer and sensor HSPA5 and other ER stress-related genes CALM2 and YKT6 indicating that ER stress response was involved in juniper berry extract mediated cell death. In composition analysis, we identified and quantified low concentrations of fifteen phenolic compounds. The main groups of them were flavones, flavonols, phenolic acids, flavanol and biflavonoid including glycosides of quercetin, apigenin, isoscutellarein and hypolaetin. It is suggested that juniper berry extract induced the p53-associated apoptosis through the potentiation and synergism by several phenolic compounds.
  • Liu, Liwei; Herfindal, Lars; Jokela, Jouni; Shishido, Tania Keiko; Wahlsten, Matti; Doskeland, Stein Ove; Sivonen, Kaarina (2014)
  • Aly, Ashraf A.; Bräse, Stefan; Hassan, Alaa A.; Mohamed, Nasr K.; Abd El-Haleem, Lamiaa E.; Nieger, Martin; Morsy, Nesrin M.; Alshammari, Mohammed B.; Ibrahim, Mahmoud A. A.; Abdelhafez, Elshimaa M. N. (2020)
    Three new series of paracyclophanyl-dihydronaphtho[2,3-d]thiazoles and paracyclophanyl-thiazolium bromides were designed, synthesized, and characterized by their spectroscopic data, along with X-ray analysis. One-dose assay results of anticancer activity indicated that 3a-e had the highest ability to inhibit the proliferation of different cancer cell lines. Moreover, the hybrids 3c-e were selected for five-dose analyses to demonstrate a broad spectrum of antitumor activity without apparent selectivity. Interestingly, series I compounds (Z)-N-substituted-4,9-dihydronaphtho[2,3-d]thiazol-3(2H)-yl)-4 '-[2.2]paracyclophanylamide) that are carrying 1,4-dihydronaphthoquinone were more active as antiproliferative agents than their naphthalene-containing congeners (series II: substituted 2-(4 '-[2.2]paracyclophanyl)hydrazinyl)-4-(naphth-2-yl)-thiazol-3-ium bromide hybrids) and (series III: 3-(4 '-[2.2]paracyclophanyl)amido-2-(cyclopropylamino)-4-(naphth-2-yl)thiazol-3-ium bromide) toward the SK-MEL-5 melanoma cell line. Further antiproliferation investigations of 3c and 3e on the healthy, normal unaffected SK-MEL-5 cell line indicated their relative safety. Compound 3c showed an inhibition of eight isoforms of cyclin-dependent kinases (CDK); however, it exhibited the lowest IC50 of 54.8 nM on CDK1 in comparison to Dinaciclib as a reference. Additionally, compound 3c revealed a remarkable downregulation of phospho-Tyr15 with a level (7.45 pg/mL) close to the reference. 3c mainly showed cell cycle arrest in the pre-G1 and G2/M phases upon analysis of the SK-MEL-5 cell line. The sequential caspase-3 assay for 3c indicated a remarkable overexpression level. Finally, a molecular docking study was adopted to elucidate the binding mode and interactions of the target compounds with CDK1.
  • Johns, Scott C.; Yin, Xin; Jeltsch, Michael; Bishop, Joseph R.; Schuksz, Manuela; El Ghazal, Roland; Wilcox-Adelman, Sarah A.; Alitalo, Kari; Fuster, Mark M. (2016)
    Rationale: Lymphatic vessel growth is mediated by major prolymphangiogenic factors, such as vascular endothelial growth factor (VEGF-C) and VEGF-D, among other endothelial effectors. Heparan sulfate is a linear polysaccharide expressed on proteoglycan core proteins on cell membranes and matrix, playing roles in angiogenesis, although little is known about any function(s) in lymphatic remodeling in vivo. Objective: To explore the genetic basis and mechanisms, whereby heparan sulfate proteoglycans mediate pathological lymphatic remodeling. Methods and Results: Lymphatic endothelial deficiency in the major heparan sulfate biosynthetic enzyme N-deacetylase/N-sulfotransferase-1 (Ndst1; involved in glycan-chain sulfation) was associated with reduced lymphangiogenesis in pathological models, including spontaneous neoplasia. Mouse mutants demonstrated tumor-associated lymphatic vessels with apoptotic nuclei. Mutant lymphatic endothelia demonstrated impaired mitogen (Erk) and survival (Akt) pathway signaling and reduced VEGF-C-mediated protection from starvation-induced apoptosis. Lymphatic endothelial-specific Ndst1 deficiency (in Ndst1(f/f)Prox1(+/CreERT2) mice) was sufficient to inhibit VEGF-C-dependent lymphangiogenesis. Lymphatic heparan sulfate deficiency reduced phosphorylation of the major lymphatic growth receptor VEGF receptor-3 in response to multiple VEGF-C species. Syndecan-4 was the dominantly expressed heparan sulfate proteoglycan in mouse lymphatic endothelia, and pathological lymphangiogenesis was impaired in Sdc4((-/-)) mice. On the lymphatic cell surface, VEGF-C induced robust association between syndecan-4 and VEGF receptor-3, which was sensitive to glycan disruption. Moreover, VEGF receptor-3 mitogen and survival signaling was reduced in the setting of Ndst1 or Sdc4 deficiency. Conclusions: These findings demonstrate the genetic importance of heparan sulfate and the major lymphatic proteoglycan syndecan-4 in pathological lymphatic remodeling. This may introduce novel future strategies to alter pathological lymphatic-vascular remodeling.
  • Almahmoudi, Rabeia; Salem, Abdelhakim; Murshid, Sakhr; Dourado, Mauricio Rocha; Apu, Ehsanul Hoque; Salo, Tuula; Al-Samadi, Ahmed (2019)
    We recently showed that extracellular interleukin-17F (IL-17F) correlates with better disease-specific survival in oral tongue squamous cell carcinoma (OTSCC) patients. However, the underlying mechanisms of such effect remain obscure. Here, we used qRT-PCR to assess the expression of IL-17F and its receptors (IL-17RA and IL-17RC) in two OTSCC cell lines (HSC-3 and SCC-25) and in normal human oral keratinocytes (HOKs). IL-17F effects on cancer cell proliferation, migration, and invasion were studied using a live-imaging IncuCyte system, and a Caspase-3/7 reagent was used for testing apoptosis. 3D tumor spheroids were utilized to assess the impact of IL-17F on invasion with or without cancer-associated fibroblasts (CAFs). Tube-formation assays were used to examine the effects of IL-17F on angiogenesis using human umbilical vein endothelial cells (HUVEC). OTSCC cells express low levels of IL-17F, IL-17RA, and IL-17RC mRNA compared with HOKs. IL-17F inhibited cell proliferation and random migration of highly invasive HSC-3 cells. CAFs promoted OTSCC invasion in tumor spheroids, whereas IL-17F eliminated such effect. IL-17F suppressed HUVEC tube formation in a dose-dependent manner. Collectively, we suggest that IL-17F counteracts the pro-tumorigenic activity in OTSCC. Due to its downregulation in tumor cells and inhibitory activity in in vitro cancer models, targeting IL-17F or its regulatory pathways could lead to promising immunotherapeutic strategies against OTSCC.
  • Erkkilä, K.; Aito, H.; Aalto, K.; Pentikäinen, V.; Dunkel, L. (2002)
    Dysregulation of male germ cell apoptosis has been associated with the pathogenesis of male infertility. Therefore, factors involved in the regulation of germ cell death are being actively investigated. Here, we studied the effects of lactate on human male germ cell death, using as a model a testis tissue culture in which physiological contacts are maintained between the germ cells and the supportive somatic Sertoli cells. Apoptosis of spermatocytes, spermatids and a few spermatogonia was induced by culturing segments of seminiferous tubules under serum-free conditions. This germ cell death was inhibited effectively and dose-dependently by lactate, indicating that it plays a crucial role in controlling cell death cascades of male germ cells. Interestingly, the anti-apoptotic role of lactate was not associated with changes in testicular adenine nucleotide (ATP, ADP and AMP) levels. In the seminiferous tubules, the final site of the death-suppressing action of lactate appeared to be downstream along the cell death pathway activated by the Fas receptor of the germ cells. In conclusion, testicular cell death was effectively regulated by lactate, which may be regarded as a potential compound for optimizing in-vitro methods involving male germ cells for assisted reproduction.
  • Lucendo, Estefania; Sancho, Monica; Lolicato, Fabio; Javanainen, Matti; Kulig, Waldemar; Leiva, Diego; Duart, Gerard; Andreu-Fernandez, Vicente; Mingarro, Ismael; Orzaez, Mar (2020)
    The Bcl-2 protein family comprises both proand antiapoptotic members that control the permeabilization of the mitochondrial outer membrane, a crucial step in the modulation of apoptosis. Recent research has demonstrated that the carboxyl-terminal transmembrane domain (TMD) of some Bcl-2 protein family mem-bers can modulate apoptosis; however, the transmembrane interactome of the antiapoptotic protein Mcl-1 remains largely unexplored. Here, we demonstrate that the Mcl-1 TMD forms homooligomers in the mitochondrial membrane, competes with full-length Mcl-1 protein with regards to its antiapoptotic function, and induces cell death in a Bok-dependent manner. While the Bok TMD oligomers locate preferentially to the endoplasmic reticulum (ER), heterooligomerization between the TMDs of Mcl-1 and Bok predominantly takes place at the mitochondrial membrane. Strikingly, the coexpression of Mcl-1 and Bok TMDs produces an increase in ER mitochondrial-associated membranes, suggesting an active role of Mcl-1 in the induced mitochondrial targeting of Bok. Finally, the introduction of Mcl-1 TMD somatic mutations detected in cancer patients alters the TMD interaction pattern to provide the Mcl-1 protein with enhanced antiapoptotic activity, thereby highlighting the clinical relevance of Mcl-1 TMD interactions.
  • Sierra-Torre, Virginia; Plaza-Zabala, Ainhoa; Bonifazi, Paolo; Abiega, Oihane; Diaz-Aparicio, Irune; Tegelberg, Saara; Lehesjoki, Anna-Elina; Valero, Jorge; Sierra, Amanda (2020)
    Objective Microglial phagocytosis of apoptotic cells is an essential component of the brain regenerative response during neurodegeneration. Whereas it is very efficient in physiological conditions, it is impaired in mouse and human mesial temporal lobe epilepsy, and now we extend our studies to a model of progressive myoclonus epilepsy type 1 in mice lacking cystatin B (CSTB). Methods We used confocal imaging and stereology-based quantification of apoptosis and phagocytosis of the hippocampus ofCstbknockout (KO) mice, an in vitro model of phagocytosis and siRNAs to acutely reduceCstbexpression, and a virtual three-dimensional (3D) model to analyze the physical relationship between apoptosis, phagocytosis, and active hippocampal neurons. Results Microglial phagocytosis was impaired in the hippocampus ofCstbKO mice at 1 month of age, when seizures arise and hippocampal atrophy begins. This impairment was not related to the lack of Cstb in microglia alone, as shown by in vitro experiments with microglial Cstb depletion. The phagocytosis impairment was also unrelated to seizures, as it was also present inCstbKO mice at postnatal day 14, before seizures begin. Importantly, phagocytosis impairment was restricted to the granule cell layer and spared the subgranular zone, where there are no active neurons. Furthermore, apoptotic cells (both phagocytosed and not phagocytosed) inCstb-deficient mice were at close proximity to active cFos(+)neurons, and a virtual 3D model demonstrated that the physical relationship between apoptotic cells and cFos(+)neurons was specific forCstbKO mice. Significance These results suggest a complex crosstalk between apoptosis, phagocytosis, and neuronal activity, hinting that local neuronal activity could be related to phagocytosis dysfunction inCstbKO mice. Overall, these data suggest that phagocytosis impairment is an early feature of hippocampal damage in epilepsy and opens novel therapeutic approaches for epileptic patients based on targeting microglial phagocytosis.
  • Haikala, Heidi M.; Klefstrom, Juha; Eilers, Martin; Wiese, Katrin E. (2016)
    Apoptosis caused by deregulated MYC expression is a prototype example of intrinsic tumor suppression. However, it is still unclear how supraphysiological MYC expression levels engage specific sets of target genes to promote apoptosis. Recently, we demonstrated that repression of SRF target genes by MYC/MIZ1 complexes limits AKT-dependent survival signaling and contributes to apoptosis induction. Here we report that supraphysiological levels of MYC repress gene sets that include markers of basal-like breast cancer cells, but not luminal cancer cells, in a MIZ1-dependent manner. Furthermore, repressed genes are part of a conserved gene signature characterizing the basal subpopulation of both murine and human mammary gland. These repressed genes play a role in epithelium and mammary gland development and overlap with genes mediating cell adhesion and extracellular matrix organization. Strikingly, acute activation of oncogenic MYC in basal mammary epithelial cells is sufficient to induce luminal cell identity markers. We propose that supraphysiological MYC expression impacts on mammary epithelial cell identity by repressing lineage-specific target genes. Such abrupt cell identity switch could interfere with adhesion-dependent survival signaling and thus promote apoptosis in pre-malignant epithelial tissue.
  • Evers, Mitchell; Ten Broeke, Toine; Jansen, J.H. Marco; Nederend, Maaike; Hamdan, Firas; Reiding, Karli R.; Meyer, Saskia; Moerer, Petra; Brinkman, Iris; Rösner, Thies; Lebbink, Robert Jan; Valerius, Thomas; Leusen, Jeanette H.W. (2020)
    ABSTRACT Current combination therapies elicit high response rates in B cell malignancies, often using CD20 antibodies as the backbone of therapy. However, many patients eventually relapse or develop progressive disease. Therefore, novel CD20 antibodies combining multiple effector mechanisms were generated. To study whether neutrophil-mediated destruction of B cell malignancies can be added to the arsenal of effector mechanisms, we chimerized a panel of five previously described murine CD20 antibodies to the human IgG1, IgA1 and IgA2 isotype. Of this panel, we assessed in vitro antibody-dependent cell-mediated cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC) and direct cell death induction capacity and studied the efficacy in two different in vivo mouse models. IgA antibodies outperformed IgG1 antibodies in neutrophil-mediated killing in vitro, both against CD20-expressing cell lines and primary patient material. In these assays, we observed loss of CD19 with both IgA and IgG antibodies. Therefore, we established a novel method to improve the assessment of B-cell depletion by CD20 antibodies by including CD24 as a stable cell marker. Subsequently, we demonstrated that only IgA antibodies were able to reduce B cell numbers in this context. Additionally, IgA antibodies showed efficacy in both an intraperitoneal tumor model with EL4 cells expressing huCD20 and in an adoptive transfer model with huCD20-expressing B cells. Taken together, we show that IgA, like IgG, can induce ADCC and CDC, but additionally triggers neutrophils to kill (malignant) B cells. We conclude that antibodies of the IgA isotype offer an attractive repertoire of effector mechanisms for the treatment of CD20-expressing malignancies.
  • Barreto, Goncalo; Manninen, Mikko; Eklund, Kari K. (2020)
    Osteoarthritis (OA) has long been viewed as a degenerative disease of cartilage, but accumulating evidence indicates that inflammation has a critical role in its pathogenesis. In particular, chondrocyte-mediated inflammatory responses triggered by the activation of innate immune receptors by alarmins (also known as danger signals) are thought to be involved. Thus, toll-like receptors (TLRs) and their signaling pathways are of particular interest. Recent reports suggest that among the TLR-induced innate immune responses, apoptosis is one of the critical events. Apoptosis is of particular importance, given that chondrocyte death is a dominant feature in OA. This review focuses on the role of TLR signaling in chondrocytes and the role of TLR activation in chondrocyte apoptosis. The functional relevance of TLR and TLR-triggered apoptosis in OA are discussed as well as their relevance as candidates for novel disease-modifying OA drugs (DMOADs).
  • Pavet, V.; Shlyakhtina, Y.; He, T.; Ceschin, D. G.; Kohonen, P.; Perala, M.; Kallioniemi, O.; Gronemeyer, H. (2014)
  • Pulkka, Olli-Pekka; Nilsson, Bengt; Sarlomo-Rikala, Maarit; Reichardt, Peter; Eriksson, Mikael; Hall, Kirsten Sundby; Wardelmann, Eva; Vehtari, Aki; Joensuu, Heikki; Sihto, Harri (2017)
    Background: The SLUG transcription factor has been linked with the KIT signalling pathway that is important for gastrointestinal stromal tumour (GIST) tumourigenesis. Its clinical significance in GIST is unknown. Methods: Influence of SLUG expression on cell proliferation and viability were investigated in GIST48 and GIST882 cell lines. The association between tumour SLUG expression in immunohistochemistry and recurrence-free survival (RFS) was studied in two clinical GIST series, one with 187 patients treated with surgery alone, and another one with 313 patients treated with surgery and adjuvant imatinib. Results: SLUG downregulation inhibited cell proliferation, induced cell death in both cell lines, and sensitised GIST882 cells to lower imatinib concentrations. SLUG was expressed in 125 (25.0%) of the 500 clinical GISTs evaluated, and expression was associated with several factors linked with unfavourable prognosis. SLUG expression was associated with unfavourable RFS both when patients were treated with surgery alone (HR = 3.40, 95% CI = 1.67-6.89, P = 0.001) and when treated with surgery plus adjuvant imatinib (HR = 1.83, 95% CI = 1.29-2.60, P = 0.001). Conclusions: GIST patients with high tumour SLUG expression have unfavourable RFS. SLUG may mediate pro-survival signalling in GISTs.
  • Shanthi Venkatesan, Aadhavan (Helsingfors universitet, 2017)
    A new myopathy called “wooden breast” affecting Pectoralis major muscles of broilers was described by researchers in Finland and other countries and it has affected the economy of the poultry industry in the last few years. The exact mechanisms behind this myopathy are yet to be discovered. This study focused on establishing any relationship between this new myodegeneration disorder “wooden breast” and small heat shock proteins. The primary aim of this work was to study the 3 important small heat shock proteins such as HSP20, HSP27 and αβ-crystallin in wooden breast and normal breast fillets by means of immunoblot technique. Six random fillet samples identified as wooden breast and six normal breast muscle samples were selected for this study. The presence of small heat shock protein HSP27 was comparatively higher in wooden breast muscles in both the myofibrillar fraction (p-value = 0. 02) and sarcoplasmic muscle fraction (p-value = 0. 01) than their presence in the normal breast muscles. Stress suffered by the birds belonging to the wooden breast condition is suggested to upregulate the heat shock proteins to antagonize against protein misfolding, protein denaturation and for protection of myofibrillar and structural proteins. As an outcome of the study, a proposal can be made about the possible mechanism behind the wooden breast to be ischemia and heat stress causing the upregulation of heat shock proteins, which was supported by the presence of HSP27 in both the myofibrillar and the sarcoplasmic muscle fraction. The presence of a relative higher amount of HSP27 in wooden breast muscles suggests that they defend against the stress suffered by the bird. Detecting the exact time point of onset of stress and factors causing the stress might unravel more information about this disorder, and studying the presence of small heat shock proteins at different time points post slaughter till the start of rigor mortis might provide additional details behind this muscle disorder.
  • Ciparyte, Auguste (Helsingin yliopisto, 2020)
    Diabetic ovarian cancer patients who take metformin as part of their anti-diabetic medication generally respond better to DNA-damaging cancer treatment. The molecular mechanisms of the anti-cancer effects of metformin are currently being investigated, but they remain poorly elucidated. Not much is understood about the metformin effect on DNA damage in ovarian cancer cells, where it is of particular importance. When chemotherapy-induced double-stranded DNA breaks are unrepaired, cells reach a point when they cannot tolerate the accumulated DNA damage and die. However, some ovarian cancer cells efficiently employ DNA repair mechanisms, the most prominent being homologous recombination (HR), to overcome DNA damage. Efficient HR causes chemoresistance. An important question is whether metformin has the ability to induce the HR-deficient state in cancer cells, thereby sensitizing them to treatment. This study did not examine HR directly, but it assessed HR indirectly by observing the effect of metformin on recovery from DNA damage in two ovarian cancer cell lines: OVCAR4 (HR-proficient) and Kuramochi (HR-deficient). Additionally, this study evaluated the metformin effect on cell proliferation and apoptosis. OVCAR4 and Kuramochi cells were exposed to varying metformin concentrations (0,5 mM, 5 mM, 10 mM, 15 mM, 20 mM and 25 mM) and for varying durations (24 hours and 48 hours). This study also tested how metformin pretreatment affected the cells’ ability to repair externally (ionizing irradiation) induced DNA damage. The cells were imaged with a high-content imaging system, and percentages of nuclei that were positive for markers for different cellular processes (i.e., DNA damage, proliferation, and apoptosis) were calculated. The study found that only high metformin concentrations, such as 20 mM were able to increase DNA damage and reduce cell proliferation in HR-proficient OVCAR4 cells, both non-irradiated and irradiated. The HR-deficient Kuramochi cell line was generally more sensitive to metformin, particularly with regards to DNA damage, which increased using metformin concentrations < 20 mM. However, 20 mM concentration resulted in the most significant effects. Similarly, only high metformin concentration (25 mM) increased apoptosis, although data were obtained only for a limited number of Kuramochi cells. More experiments on apoptosis would be beneficial. Also, more extensive experiments for the irradiation part are needed to validate these preliminary findings, as well as examining whether high metformin concentrations (> 20 mM) affect specifically the HR-mediated DNA repair pathway.
  • Kilpeläinen, Niko (Helsingfors universitet, 2014)
    Type 1 diabetes (T1D) is a chronic autoimmune disease in which insulin-producing beta-cells in pancreatic islets are destroyed by the body's own immune system. Patients with T1D require a life-long treatment with exogenous insulin. In Finland the incidence of T1D is the highest in the world. Interleukin-17 producing T helper cells (Th17 cells) have been linked to the disease pathogenesis in animal models of T1D. Increased activation of Th17-immunity, including elevated expression of IL-17 and interleukin-22 (IL-22) in the peripheral blood has been associated with human T1D. Additionally, IL-17 has been demonstrated to be harmful to mouse beta cells and human pancreatic islets. The publications reporting detrimental effects of IL-17 on mouse beta cells and human pancreatic islets lack the information regarding the role of IL-22 in the inflammatory conditions mediated by the pro-inflammatory cytokines IL-1b, IFN-? and IL-17. The aim of this study was to examine the effects of IL-17 and IL-22 on mouse beta cells and human pancreatic islets under the inflammatory conditions mediated by IL-1b and IFN-?. Cytokine-induced apoptosis of mouse beta cells was studied using MIN6 cells. Gene expression of anti-apoptosis, beta-cell function and IL-17-signaling related genes was studied from cytokine-stimulated MIN6 cells. Stress response and apoptosis related genes were studied from the human pancreatic islets. The proportion of apoptotic and necrotic cells of cytokine stimulated MIN6 cells was studied with fluorescence microscopy. Gene expression analyses were performed with quantitative reverse transcription qPCR (RT-qPCR). In this study IL-17 and IL-22 was demonstrated to modulate the outcome of IL-1b and IFN-? induced stress response in mouse beta-cells and human pancreatic islets as measured by the changes in the frequency of apoptotic cells and gene expression of stress, function and cytokine-signaling related genes.
  • Id, Linda (Helsingin yliopisto, 2022)
    Breast cancer is the most common cancer in the world and among women the most cancer deaths causing cancer. MYC is a proto-oncogene, which becomes oncogenic when its expression is deregulated in cancer. MYC is commonly overexpressed in human tumours and this alteration is associated with aggressive cancer phenotype. Furthermore, alterations in the MYC network have been found in the great majority of breast cancers. MYC promotes mitochondrial apoptosis causing a cancer vulnerability, however, in cancer cells the apoptosis is often prevented by antiapoptotic BCL-2 family members. In this study, cell viability and cell death analysis of treated triple-negative breast cancer cell lines together with dendritic cell activation experiments were conducted. This study aimed to find the most potent BCL-2 family antagonist (BH3 mimetic) to combine with metformin to overcome the antiapoptotic BCL-2 family proteins inhibition of MYC-induced apoptosis. In addition, this study determined whether the combinations could induce immunogenic cell death to further intensify cancer cell killing through anti-tumour immunity. In this study, BH3 mimetics combined with metformin were found to induce cell death and reduce cell viability in TNBC cell lines. In addition, metformin and BH3 mimetics were found to activate dendritic cells directly and through immunogenic cell death of cancer cells. However, no MYC-dependent cell death or immunogenic cell death were observed, and this study was unable to indicate the most potent BH3 mimetic to combine with metformin.