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1.
Int J Mol Sci ; 24(22)2023 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-38003481

RESUMO

Dormant primordial follicles (PMF), which constitute the ovarian reserve, are recruited continuously into the cohort of growing follicles in the ovary throughout female reproductive life. Gonadotoxic chemotherapy was shown to diminish the ovarian reserve pool, to destroy growing follicle population, and to cause premature ovarian insufficiency (POI). Three primary mechanisms have been proposed to account for this chemotherapy-induced PMF depletion: either indirectly via over-recruitment of PMF, by stromal damage, or through direct toxicity effects on PMF. Preventative pharmacological agents intervening in these ovotoxic mechanisms may be ideal candidates for fertility preservation (FP). This manuscript reviews the mechanisms that disrupt follicle dormancy causing depletion of the ovarian reserve. It describes the most widely studied experimental inhibitors that have been deployed in attempts to counteract these affects and prevent follicle depletion.


Assuntos
Antineoplásicos , Reserva Ovariana , Humanos , Feminino , Folículo Ovariano , Ovário , Apoptose , Antineoplásicos/efeitos adversos
2.
Hum Reprod ; 38(6): 1086-1098, 2023 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-37015102

RESUMO

STUDY QUESTION: To what extent and how does combined administration of the follicle activation pathway suppressive agents temsirolimus (Tem) and c-terminus recombinant anti-Müllerian hormone (rAMH) protect against chemotherapy-induced ovarian reserve loss? SUMMARY ANSWER: Combined administration of Tem and rAMH completely prevents cyclophosphamide (Cy)-induced follicle depletion and protects the ovarian reserve in mice, primarily via primordial follicle (PMF) suppression of activation and to a lesser degree by reducing apoptosis. WHAT IS KNOWN ALREADY: There is conflicting evidence regarding the contributory roles of apoptosis and follicle activation in chemotherapy-induced PMF loss. Tem, a mammalian target of rapamycin (mTOR) inhibitor, reduces activity of the phosphoinositide 3-kinases-phosphatase and tensin homolog (PI3K-PTEN) pathway which provides intrinsic regulation of PMF activation. Anti-Müllerian hormone (AMH), secreted by early growing follicles, is an extrinsic regulator of PMF activation. STUDY DESIGN, SIZE, DURATION: Whole ovaries of 12-day-old mice were cultured ex vivo for 7 days in the presence of Cy ± rAMH or Tem. Eight-week-old mice were randomized into eight treatment groups: vehicle control/rAMH/Tem/Cy/Tem + rAMH/Cy + Tem/Cy + rAMH/Cy + Tem + rAMH. Twelve hours after treatment, ovaries were removed for DNA damage analysis, and 24 h after treatment either for analysis of PI3K pathway proteins or to be fixed and immunostained for analyses of proliferation and apoptosis. Three or 21 days following treatment, ovaries were fixed and sectioned for follicle counting. PARTICIPANTS/MATERIALS, SETTING, METHODS: Hematoxylin and eosin staining was used for differential follicle counts of primordial, primary, and secondary follicles in ex vivo (n = 16-18 ovaries per group) and in vivo ovaries (n = 8 mice per group). Histological analyses were carried out to measure proliferation by quantifying Ki-67-positive granulosa cells in primary follicles (n = 4 mice per group). DNA damage and apoptosis were measured by quantification of phosphorylated form of histone 2AX (γH2AX) and cleaved poly (ADP-ribose) polymerase (cPARP)-positive PMF oocytes, respectively (n = 8 mice per group). Protein extracts from whole ovaries were analyzed by western blotting. MAIN RESULTS AND THE ROLE OF CHANCE: In vivo experiments show that treatment with Cy alone caused significant loss of PMF reserve (32 ± 2.12 versus 144 ± 2.8 in control, P < 0.001), and this was significantly attenuated by treatment with either Tem (P < 0.001) or rAMH (P < 0.001). Combined cotreatment with Cy + Tem + rAMH provided complete protection of the PMF reserve, with no significant difference in numbers of PMF versus untreated animals. Similar results were demonstrated in the ex vivo experiments. Proliferation marker Ki-67 staining was significantly reduced in granulosa cells of primary follicles in the Cy + Tem + rAMH group compared with Cy alone group (after 24 h in vivo administration of Cy, 16% versus 65%, respectively; P < 0.001). Protein analysis demonstrated not significant increased phosphorylation of follicle activation proteins rpS6 and mTOR with in vivo administration of Cy alone (1.9 and 1.4 times the control ovaries, respectively), and this was reduced to below control levels in the Cy + Tem + rAMH group (P < 0.01). The Cy + Tem + rAMH combined cotreatment protected the follicle reservoir via inhibition of Cy-induced upregulation of the PI3K signaling pathway, together with replacement of AMH suppression of PMF activation with rAMH, implying a complementary effect of the two inhibitors. The DNA damage marker γH2AX was highly positive in PMF oocytes from Cy-treated ovaries 12 h after treatment, compared with controls (94% versus 59%, respectively, P < 0.001) and was significantly reduced to (69%) in Cy + Tem + rAMH cotreated ovaries (P < 0.001). However, only 22% of PMF oocytes of the Cy group showed apoptosis at 24 h, and this was significantly reduced (12%) in ovaries after treatment with Cy + Tem + rAMH (P < 0.01). This suggests that it is not possible to equate DNA damage with oocyte death, and also indicates that less than one-third of the total PMF loss can be attributed to apoptosis, implying that most of the PMF depletion results from PMF activation but that both mechanisms play a significant role. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: The experimental design was limited by the selection of one time point for analysis of PMF activation and apoptosis (i.e. 24 h after Cy administration), although DNA damage was measured at 12 h after Cy administration and any impact on short-term follicle dynamics at 3 days after treatment. Protein analysis was conducted on whole ovary lysates therefore the protein changes identified cannot be localized to specific cells within the ovary. However, this complementary assay showed that there was activation in the ovary through massive reduction in the phosphorylation of key proteins in the PI3K cascade (rpS6 and mTOR), which is consistent with the sequence of events after Cy administration. WIDER IMPLICATIONS OF THE FINDINGS: Understanding the complementary nature of different follicle activation pathways and the impact of their suppression in prevention of chemotherapy-induced ovotoxic damage, as well as their involvement in DNA damage inhibition, provides an interesting direction for future research, and the potential for noninvasive pharmacological fertility preservation. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by a grant from the Morris Kahn Foundation. The authors declare no conflicts of interest.


Assuntos
Antineoplásicos , Reserva Ovariana , Feminino , Animais , Camundongos , Reserva Ovariana/fisiologia , Hormônio Antimülleriano/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Antígeno Ki-67 , Ciclofosfamida , Serina-Treonina Quinases TOR , Mamíferos/metabolismo
3.
Lupus ; 30(5): 775-784, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33554716

RESUMO

INTRODUCTION: Antiphospholipid syndrome (APS) is an autoimmune disorder manifested by thromboembolic events, recurrent spontaneous abortions and elevated titers of circulating antiphospholipid antibodies. In addition, the presence of antiphospholipid antibodies seems to confer a fivefold higher risk for stroke or transient ischemic attack. Although the major antigen of APS is ß2 glycoprotein I, it is now well established that antiphospholipid antibodies are heterogeneous and bind to various targets. Recently, antibodies to Annexin A2 (ANXA2) have been reported in APS. This is of special interest since data indicated ANXA2 as a key player in fibrinolysis. Therefore, in the present study we assessed whether anti-ANXA2 antibodies play a pathological role in thrombosis associated disease. MATERIALS AND METHODS: Mice were induced to produce anti-ANXA2 antibodies by immunization with ANXA2 (iANXA2) and control mice were immunized with adjuvant only. A middle cerebral artery occlusion stroke model was applied to the mice. The outcome of stroke severity was assessed and compared between the two groups. RESULTS: Our results indicate that antibodies to ANXA2 lead to a more severe stroke as demonstrated by a significant larger stroke infarct volume (iANXA2 133.9 ± 3.3 mm3 and control 113.7 ± 7.4 mm3; p = 0.017) and a more severe neurological outcome (iANXA2 2.2 ± 0.2, and control 1.5 ± 0.18; p = 0.03). CONCLUSIONS: This study supports the hypothesis that auto-antibodies to ANXA2 are an independent risk factor for cerebral thrombosis. Consequently, we propose screening for anti-ANXA2 antibodies should be more widely used and patients that exhibit the manifestations of APS should be closely monitored by physicians.


Assuntos
Anexina A2/imunologia , Síndrome Antifosfolipídica/imunologia , Autoanticorpos/imunologia , Trombose Intracraniana/metabolismo , Adulto , Animais , Anexina A2/administração & dosagem , Anexina A2/metabolismo , Anticorpos Antifosfolipídeos/sangue , Síndrome Antifosfolipídica/complicações , Síndrome Antifosfolipídica/metabolismo , Autoanticorpos/metabolismo , Autoimunidade/imunologia , Modelos Animais de Doenças , Feminino , Fibrinólise/imunologia , Humanos , Infarto da Artéria Cerebral Média/imunologia , Infarto da Artéria Cerebral Média/fisiopatologia , Injeções Subcutâneas , Trombose Intracraniana/etiologia , Ataque Isquêmico Transitório/imunologia , Camundongos , Camundongos Endogâmicos BALB C/imunologia , Pessoa de Meia-Idade , Fatores de Risco , Índice de Gravidade de Doença , Acidente Vascular Cerebral/imunologia , beta 2-Glicoproteína I/metabolismo
4.
Fertil Steril ; 105(1): 20-9, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26677788

RESUMO

Recent advances in our understanding of the mechanisms underlying the impact of cytotoxic drugs on the ovary have opened up new directions for the protection of the ovary from chemotherapy-induced damage. These advances have spurred the investigation of pharmacological agents to prevent ovarian damage at the time of treatment. Prevention of ovarian damage and follicle loss would provide significant advantages over existing fertility preservation techniques. This manuscript reviews new methods for the prevention of chemotherapy-induced ovarian damage, including agents that act on the PI3K/PTEN/Akt follicle activation pathway, apoptotic pathways, the vascular system, and other potential methods of reducing chemotherapy-induced ovotoxicity.


Assuntos
Antineoplásicos/efeitos adversos , Fertilidade/efeitos dos fármacos , Infertilidade Feminina/prevenção & controle , Ovário/efeitos dos fármacos , Substâncias Protetoras/farmacologia , Animais , Antineoplásicos/metabolismo , Apoptose/efeitos dos fármacos , Citoproteção , Desenho de Fármacos , Feminino , Humanos , Infertilidade Feminina/induzido quimicamente , Infertilidade Feminina/metabolismo , Infertilidade Feminina/patologia , Infertilidade Feminina/fisiopatologia , Terapia de Alvo Molecular , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/metabolismo , Folículo Ovariano/patologia , Ovário/metabolismo , Ovário/patologia , Ovário/fisiopatologia , Transdução de Sinais/efeitos dos fármacos
5.
Eukaryot Cell ; 9(9): 1403-15, 2010 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-20656913

RESUMO

cspA (for cell surface protein A) encodes a repeat-rich glycophosphatidylinositol (GPI)-anchored cell wall protein (CWP) in the pathogenic fungus Aspergillus fumigatus. The number of repeats in cspA varies among isolates, and this trait is used for typing closely related strains of A. fumigatus. We have previously shown that deletion of cspA is associated with rapid conidial germination and reduced adhesion of dormant conidia. Here we show that cspA can be extracted with hydrofluoric acid (HF) from the cell wall, suggesting that it is a GPI-anchored CWP. The cspA-encoded CWP is unmasked during conidial germination and is surface expressed during hyphal growth. Deletion of cspA results in weakening of the conidial cell wall, whereas its overexpression increases conidial resistance to cell wall-degrading enzymes and inhibits conidial germination. Double mutant analysis indicates that cspA functionally interacts with the cell wall protein-encoding genes ECM33 and GEL2. Deletion of cspA together with ECM33 or GEL2 results in strongly reduced conidial adhesion, increased disorganization of the conidial cell wall, and exposure of the underlying layers of chitin and beta-glucan. This is correlated with increasing susceptibility of the DeltacspA, DeltaECM33, and DeltacspA DeltaECM33 mutants to conidial phagocytosis and killing by human macrophages and hyphal damage induced by neutrophils. However, these strains did not exhibit altered virulence in mice with infected lungs. Collectively, these results suggest a role for cspA in maintaining the strength and integrity of the cell wall.


Assuntos
Aspergilose/microbiologia , Aspergillus fumigatus/metabolismo , Parede Celular/metabolismo , Proteínas Fúngicas/metabolismo , Interações Hospedeiro-Patógeno , Esporos Fúngicos/metabolismo , Animais , Aspergillus fumigatus/genética , Aspergillus fumigatus/crescimento & desenvolvimento , Aspergillus fumigatus/patogenicidade , Linhagem Celular , Parede Celular/genética , Feminino , Proteínas Fúngicas/genética , Regulação Fúngica da Expressão Gênica , Humanos , Camundongos , Camundongos Endogâmicos ICR , Ligação Proteica , Esporos Fúngicos/genética , Esporos Fúngicos/crescimento & desenvolvimento , Esporos Fúngicos/patogenicidade , Virulência
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