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"LATAR BELAKANG : Salah satu hambatan dalam program reproduksi berbantu adalah rendahnya viabilitas dan motilitas sel spermatozoa. Analisis proteomik menunjukkan adanya banyak protein yang diduga berperan dalam regulasi motilitas dan viabilitas spermatozoa antara lain progesteron. Penelitian ini bertujuan untuk menganalisis efek prosurvival progesteron terhadap spermatozoa melalui penekanan apoptosis.BAHAN DAN CARA KERJA : Sampel spermatozoa dicuci dengan sentrifugasi gradient. Sampel spermatozoa di tambahkan progesteron dengan konsentrasi 0 kontrol , 250, 500, 750 dan 1000 ng/mL. Setelah perlakuan, sampel dilakukan pemeriksaan integritas membran dengan metode HOS dan pemeriksaan motilitas dengan Computer Assisted Sperm Analyzer CASA . Deteksi protein fosforilasi tirosin dan Akt serta aktivitas kaspase dilakukan dengan metode western blot.HASIL : Efek penambahan progesteron meningkatkan rerata motilitas spermatozoa namun berbeda tidak bermakna p>0.05 . Integritas membran spermatozoa tidak berpengaruh pada pemberian progesteron. Analisis western blot menunjukkan peningkatan fosforilasi protein tirosin antara kelompok kontrol dan setelah diberikan progesteron p>0.05 . Demikian halnya dengan hasil fosforilasi protein Akt juga mengalami peningkatan pada kelompok kontrol dan setelah diberikan progesteron berbagai dosis. Aktivitas kaspase-3 mengalami penurunan bila dibandingkan antara kelompok kontrol dan setelah diberikan progesteron p

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BACKGROUND One of the obstacles in assisted reproduction programs is the low viability and motility of spermatozoa cells. Proteomic analysis indicates that many proteins are thought to play a role in the regulation of motility and viability of spermatozoa, among others, progesterone. This study aims to analyze the prosurvival effect of progesterone against spermatozoa through apoptosis suppression.METHODS Spermatozoa was washed with gradient centrifugation. Progesterone is added to each sample with a final concentration 0 control , 250, 500, 750 and 1000 ng mL. After the sample treatment was done, membrane integrity checking with hypoosmotic swelling test and motility examination with Computer Assisted Sperm Analyzer CASA . Detection of protein in the western blot will be done that recognizes the phosphorylation of tyrosine residues and Akt and caspase activity.RESULT The effect of addition of progesterone increases sperm motility but not signicantly different p 0.05 . The integrity of the spermatozoa membrane is no effect in progesterone. Western blot analysis revealed an increase of tyrosine phosphorylation protein levels between control and after progesterone group p 0.05 . Similarly, the results of Akt protein phosphorylation also increased in control and after progesterone group. Caspase 3 activity decreased when compared between control and after progesterone group p "

"Apoptosis or programmed cell death is a normal condition for development and live multicellular organism. Apoptosis is a morphological phenomenon that play important role in physiological processes during fetal development and in adult. Mitochondria play an important role in apoptosis. Mitochondria can do apoptosis directly. Mitochondria has 2 family of protein Bcl-2. Bcl-2 and Bcl-XL are anti apoptosis while Bad and Bax are pro apoptosis. There are 3 different mechanism apoptosis. One generated by signals arising within the cell another triggered by death activators binding to receptors at the cell surface and a third may be triggered by dangerous agent that different from two ways before. Apoptosis also need caspase as cell death executor. Study of apoptosis still done especially in case of disease. Some disease have known related with disturbing of apoptosis mechanism for example cancer and auto immun. This article reviews about molecular mechanism of apoptosis for understanding disease and future therapy."

"Sel sperma manusia memproduksi reactive oxygen species (ROS) selama respirasi mitokondria dalam jumlah rendah yang dapat membantu berbagai jalur persinyalan. Produksi ROS fisiologis pada sel sperma dapat mengatur karakteristik fungsional yang penting seperti motilitas, kapasitasi, reaksi akrosom, hiperaktivasi, dan fusi sperma-oosit. Namun ROS yang terlalu banyak justru akan menyebabkan efek sebaliknya. Pada penelitian sebelumnya, dilaporkan bahwa α-tokoferol mampu meningkatkan motilitas dan melindungi sperma dari efek buruk stres oksidatif. Namun mekanisme molekuler efek tersebut masih belum jelas. Pada penelitian ini, dilakukan suplementasi α-tokoferol pada sel sperma untuk dianalisis terhadap beberapa parameter diantaranya, kadar MDA, motilitas, integritas membran, kapasitasi melalui ekspresi fosforilasi tirosin, ketahanan hidup melalui ekspresi Akt pada sel sperma, dan apoptosis melalui ekspresi caspase 3. Hasil dari penelitian ini menujukkan bahwa penambahan α-tokoferol tidak dapat menurunkan kadar MDA sel sperma. Namun pada parameter lain, penambahan α-tokoferol dapat meningkatkan motilitas, integritas membran sel, ekspresi fosforilasi tirosin, ekspresi fosforilasi Akt, dan menurunkan ekspresi caspase 3 pada sel sperma.

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The sperm cells of humans produce reactive oxygen species (ROS) during mitochondrial respiration in low amounts that can aid various signaling pathways. Physiological ROS production in sperm cells can regulate important functional characteristics such as motility, capacitation, acrosome reaction, hyperactivation, and sperm-oocyte fusion. However, an excess of ROS can have adverse effects. In previous studies, it has been reported that α-tocopherol can enhance motility and protect sperm from the harmful effects of oxidative stress. However, the molecular mechanisms of these effects are still unclear. In this study, α-tocopherol supplementation was performed on sperm cells to analyze several parameters, including MDA levels, motility, membrane integrity, capacitation through tyrosine phosphorylation expression, survival through Akt expression in sperm cells, and apoptosis through caspase 3 expression. The results of this study indicate that the addition of α-tocopherol cannot reduce MDA levels in sperm cells. However, in the other parameters, the addition of α-tocopherol can increase motility, membrane integrity, tyrosine phosphorylation expression, Akt phosphorylation expression, and decrease caspase 3 expression in sperm cells."

"Pendahuluan: Ekspresi berlebih suatu molekul bernama survivin dapat mencegah terjadinya apoptosis dengan menghambat caspase 9 dan mempertahankan keganasan pada sel glioblastoma. Penggunaan secretome sel punca tali pusat dapat mencegah perkembangan glioblastoma meskipun penggunaanya masih kontroversial. Untuk menguji efektifitas apoptosis secretome sel punca tali pusat pada glioblastoma, dapat dilakukan pengukuran survivin dan caspase 9 yang merupakan komponen penting pada jalur apoptosis intrinsik. Dengan demikian penelitian ini bertujuan untuk menganalisis pengaruh sekretom sel punca tali pusat terhadap ekspresi survivin dan caspase9 sebagai molekul yang mempengaruhi pertumbuhan glioblastoma Metode: Desain eksperimental in vitro dengan menggunakan galur sel glioblastoma T98G. Conditioned medium (CM) yang mengandung secretome sel punca tali pusat diperoleh dari medium sel punca  tali pusat yang dikultur selama 24 jam dengan  Î±MEM yang tidak mengandung serum. CM diencerkan 2 kali  dengan  medium tinggi glukosa Dulbecco's Modified Eagle Medium (DMEM) (50% konsentrasi). Perlakuan Sel glioblastoma T98G yang diberikan CM 50% dilakukan selama 24 jam, kemudian dilakukan ekstraksi RNA total dari sel T98G tersebut. RNA total digunakan untuk analisis ekspresi gen dengan menggunakan real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). Metode Livak dilakukan untuk menghitung ekspresi relatif  caspase 9 dan survivin dengan 18srRNA sebagai gen referensi.

Hasil: Ekspresi mRNA survivin meningkat 3.5 kali (p=0.002) dan ekspresi mRNA caspase 9 meningkat 1.6 kali (p=0.017) pada sel T98G yang diberikan CM50% dibandingkan sampel kontrol. Kesimpulan : CM  sel punca pada tali pusat meningkatkan ekspresi survivin dan caspase 9 pada sel glioblastoma T98G. Penelitian selanjutnya diperlukan untuk mengetahui pengaruh peningkatan ekspresi gen tersebut terhadap proliferasi sel glioblastoma serta mekanisme molekulernya.

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Introduction: Aberrant expression of a molecule called survivin has been suspected to prevent apoptosis by indirectly inhibits a critical apoptosis enzyme called caspase 9, maintaining tumorigenicity of glioblastoma. Umbilical cord mesenchymal stem cells (UCMSC) has been discovered to inhibit glioblastoma growth but its usage is still controversial. To measure the apoptosis effectiveness of UCMSC-CM secretome against glioblastoma, measuring survivin and caspase 9, as essential components in intrinsic apoptosis pathway, are required. Therefore, this research aims to analyze the effect of UCMSC-CM against survivin and caspase 9 expression as the molecules that affect the growth of glioblastoma.

Method: In vitro experimental design by using glioblastoma cell line T98G. Conditioned medium (CM) which contain umbilical cord mesenchymal stem cell (UCMSC) secretome was obtained from UCMSC-CM that was cultured for 24 hours with αMEM that does not contain serum. CM was diluted twice with high glucose Dulbecco’s Modified Eagle Medium (DMEM) (50% concentration). The treatment of T98G glioblastoma cell, that had been exposed to 50% CM, was performed for 24 hours, then total RNA extraction was carried out from the T98G cells. Total RNA was used to analyze genetic expression by using real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). Livak’s method was used to calculate relative expression of Survivin and Caspase 9 expression with 18srRNA as reference gene.

Results: Survivin mRNA expression increased 3.5-fold (p=0.002) while caspase 9 mRNA expression increased 1.6-fold (p=0.017) in T98G cell that was given CM50% compared with control sample.

Conclusion: UCMSC-CM increases survivin and caspase 9 expression in glioblastoma T98G cells. Future researches are required to identify the effect of the elevated genetic expressions against glioblastoma cell proliferation as well as their molecular mechanisms."

"Latar Belakang: Hiperglikemia akut maupun kronis dapat menyebabkanperubahan patalogis pada lapisan serabut saraf retina RNFL . Pada tahap dinilapisan sel ganglion GCs mengalami apoptosis, yang diregulasi oleh proteinBrn3b. Proses ini diketahui terjadi lebih awal sebelum perubahan histopatologispada RNFL terjadi pada diabetes. Proses apoptosis ditandai dengan, peningkatanstres oksidatif, peningkatan NO, NF-??, TNF-?, dan Caspase 3 pada patogenesisglaukoma primer sudut terbuka GPSTa . Asumsi klinis saat ini bahwa penderitadiabetes melitus dalam jangka waktu lama lebih dari 5 tahun kemungkinan besarberisiko terjadinya glaukoma. Namun kenyataannya dalam pengalaman klinistidak semua pasien diabetes melitus dapat terjadi GPSTa. Berdasarkan temuanklinis tersebut diduga ada peran faktor genetik yang mendasari etiologi dariGPSTa pada pasien diabetes melitus. Tujuan: Mengetahui peran gen Brn3b pada apoptosis di RGCs dan kaitannyadengan perubahan kuantitas NO, Caspase 3, NF-?B, dan TNF-? sebagai prediksiglaukoma dini pada tikus diabetes. Metode: Penelitian ekperimental in vivo menggunakan tikus jantan SpraqueDawleyusia ge; 2 bulan dengan berat 150-200 gram diambil secara random sejakNopember 2015 hingga Juli 2016. Hewan coba dibagi menjadi 2 kelompok yaitu: kelompok perlakuan diinjeksi intraperitoneal STZ 50 mg/kg dalam 0.01M buffersitrat dalam ph 4.5 dan kelompok kontrol tidak ada perlakuan. Glukosa darahpuasa diperiksa 3 hari setelah injeksi STZ, dan dikonfirmasi ge; 250 mg/dL.Jaringan retina dibagi menjadi dua bagian pada kelompok perlakuan maupunkontrol yakni retina kanan untuk IHK Caspase 3 dan TNF-? dan retina kiridibagi dua bagian yaitu untuk pemeriksaan quantitative Real-time PCR RNAdiekstraksi untuk analisis ekspresi gen Brn3b dan pemeriksaan ELISA NO danNF-?B. Hasil: Terjadi penurunan ekpresi gen Brn3b pada tikus perlakuandibandingkan ekspresi gen Brn3b kontrol sebesar 1.2677 kali bulan kedua, 1.1348kali bulan keempat, dan 2.4600 kali bulan keenam. Disisi lain terjadi penurunankuantitas NO dan peningkatan kuantitas Caspase 3, NF-?B, dan TNF-?. Kesimpulan: Ekspresi mRNA Brn3b berbanding terbalik dengan apoptosis padaRGCs. Kuantitas NO, Caspase 3, NF-?B, dan TNF-? dipengaruhi oleh ekspresiBrn3b pada RGCs akibat hiperglikemia pada tikus diabetes.

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Background: Acute and chronic hyperglycemia may pathologically changeretinal nerve fiber layer RNFL . At early stage, ganglion cells GCs undergoapoptosis, which is regulated by a Brn3b protein. This change is known to occurat earlier time before retinal histological changes can be detected in diabeticpatients. The apoptosis process is marked by an increase in oxidative stress, a risein NO, NF-??, caspase 3, and TNF-? levels in pathogenesis of primary open angleglaucoma POAG . Current clinical assumption proposes that individualssuffering from diabetes mellitus for more than 5 years have greater risk ofglaucoma. Nonetheless, clinical experience shows that not all diabetic patientsdevelop POAG. Based on clinical examinations it is suspected that there is agenetic factor that caused the etiology of POAG in diabetes mellitus patients.

Purpose: To learn the role of Brn3b gene in apoptosis of RGCs and its effect onthe changing of quantity of NO, Caspase 3, NF-?B, and TNF-? as early predictorof glaucoma in diabetic rats.

Methods: Experimental in-vivo study was carried out using male SpraqueDawleyrats with age ge; 2 months, weighing 150-200 grams. The rats wererandomly selected from November 2015 to July 2016. The animals were dividedinto two groups. Group receiving intraperitoneal injection of STZ 50 mg/kg in0.01M citric buffer and pH 4,5; 2 and control group with no treatment. Fastingblood glucose was checked 3 days after injection of STZ and hyperglycemia wasdetermined as fasting blood glucose ge; 250 mg/dL. Retinal tissue was divided intotwo parts both experimental and control groups respectively : a right retina forIHC Caspase 3 and TNF-? ; b left retina was divided into two parts for thepurpose of quantitative Real-Time PCR test RNA extraction for Brn3b geneexpression analysis and ELISA test NO and NF-?B.

Results: Experimental group showed a decrease in Brn3b expression compared tocontrol group 1.2677-fold lower on 2nd month; 1.1348-fold lower on 4th monthand 2.4600-fold lower on 6th month . On the other hand, there was a decrease ofNO and there was increased of Caspase 3, NF-?B and TNF-? quantity.

Conclusion: The expression of mRNA Brn3b is inversely proportional toapoptosis in RGCs. The quantity of NO, Caspase 3, NF-?B and TNF-? isinfluenced by expression of Brn3b in RGCs caused by hyperglycemia in diabeticrats."

"GnRH digunakan dalam program fertilisasi in vitro (FIV) sebagai salah satu regimen stimulasi ovarium. Agonis GnRH memiliki efek langsung maupun tidak langsung terhadap perkembangan endometrium terutama saat fase implantasi. Penggunaan agonis GnRH dapat berefek negatif terhadap perkembangan endometrium setelah pemberian stimulasi ovarium terhadap ekspresi reseptor dan apoptosis sel endometrium. Tujuan dari penelitian ini adalah menganalisis dampak pemberian agonis GnRH terhadap ekspresi reseptor GnRH dan protein apoptosis sel-sel endometrium fase luteal terhadap perkembangan endometrium. Sampel dari penelitian ini menggunakan bahan biologi tersimpan berupa serum dan jaringan endometrium Macaca nemestrina. Total sampel ada 8 yang terbagi atas 2 kelompok, Stimulasi dan Kontrol. Setiap sampel dilakukan 2 pemeriksaan yaitu Enzym-Linked Immunosorebent Assay (ELISA) untuk serum dan Imunohistokimia (IHK) untuk jaringan endometrium. Jaringan IHK diperiksa dengan 2 jenis antibody, reseptor GnRH dan Caspase 3. Consentrasi diukur menggunakan ELISA reader lalu dikonversi dengan Optical Density (OD) menggunakan software SoftMax Pro. Sel pada jaringan IHK dihitung secara kuantitatif berdasarkan pewarnaan menggunakan software ImageJ lalu dinilai menggunakan IHC Optical Density Score. Tidak ada perbedaan signifikan pada serum GnRH, Reseptor GnRH, dan Caspse 3 diantara kedua kelompok (p> 0,05). Terdapat korelasi negatif pada serum GnRH dengan reseptor GnRH (p=0,014; r=-0,762). Tidak terdapat korelasi antara serum GnRH dengan caspase 3 (p>0,05). Tidak ada korelasi antara reseptor GnRH dengan caspase 3 (p>0,05).

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GnRH is widely used in the embryo fertilization program as one of the ovarian stimulation regimens. At the implantation window, GnRH agonists are known to have an effect on the endometrium directly or indirectly. GnRH estimated has a negative effect on the development of endometrial cells after ovarian stimulation. This study is to analyze the impact of GnRH agonist on ovarian stimulation procedures on receptor expression and endometrial cell apoptosis due to endometrial development. The study sample was a stored biological material (BBT) from serum and the endometrial tissue of Macaca nemestrina. The total sample is 8 and divided into 2 groups, the stimulated and control groups. Each sample will be examined 2 types which are the enzyme-linked immunosorbent assay (ELISA) for serum and immunohistochemistry (IHC) for endometrial tissue. IHC was performed with anti-GnRH receptor and caspase 3 antibody. Serum concentration is measured using an ELISA reader and then converts to a concentration using SoftMax Pro Software. Quantitative data of IHC were performed using the Image-J Analyzer programs and scored by IHC Optical Density Score. There is no significant difference between GnRH serum, GnRH receptors, and Caspase 3 in stimulation or control group (p>0,05). There was a strong negative correlation between serum GnRH levels and GnRH receptors (p=0,14; r=-0,762). There was no correlation between GnRH in serum with activation of caspase 3 (p>0,05). There was no correlation between GnRH receptors with activation of caspase 3 (p>0,05)."

"Telah dilakukan penelitian untuk mengidentifikasi efek krioprotektif madu lengkeng (Dimocarpus longan) terhadap integritas struktur folikel preantral dan profil ekspresi protein apoptosis jalur intrinsik. Sebanyak 24 ekor tikus (Rattus norvegicus L) dikelompokkan menjadi 8 kelompok, terdiri atas KKN, KKV (NaCl 0,9%), KKP1 (EG 7,5%), KKP2 (EG 15%), KP1 (EG 7,5% + ML 7,5%), KP2 (EG 7,5% + ML 15%), KP3 (EG 15% + ML 7,5%), dan KP4 (EG 15% + ML 15%). Pengamatan terhadap densitas,struktur folikel serta ekspresi protein Bax,Bcl2 dan Caspase3 dilakukan terhadap sayatan ovarium yang dibuat dengan metode parafin dengan pewarnaan Hematoksilin-Eosin (HE) dan imunohistokimia. Antibodi primer yang digunakan adalah antibodi poliklonal Rabbit Anti-Bax (A00183 Boster, USA), Rabbit Anti-Bcl-2 (A00040-2 Boster, USA) dan Active Caspase-3 Rabbit Polyclonal Antibody (ab4051 Abcam, UK) dan One Step Neopoly Detection System Kit (BGNK-0025 Biogear, USA). Identifikasi terhadap tiap tipe folikel preantral menggunakan mikroskop cahaya yang terhubung dengan perangkat lunak Image Raster dan IHC profiler. Hasil penelitian menunjukkan, efek krioprotektif madu lengkeng dapat meningkatkan densitas folikel, indeks folikel intak G2 dan G3, menurunkan indeks folikel G1, menekan ekspresi protein Bax dan caspase 3 serta meningkatkan ekspresi protein Bcl2. Dengan demikian, madu lengkeng memiliki potensi untuk dikembangkan sebagai krioprotektan ekstraselular alami dalam aplikasi vitrifikasi ovarium.

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A study was conducted to identify the cryoprotective effect of longan honey (Dimocarpus longan) on the structural integrity of the preantral follicle and the expression profile of the intrinsic pathway of apoptosis protein. A total of 24 rats (Rattus norvegicus L) were grouped into 8 groups, consisting of KKN, KKV (NaCl 0.9%), KKP1 (EG 7.5%), KKP2 (EG 15%), KP1 (EG 7.5% + LH 7.5%), KP2 (EG 7.5% + LH 15%), KP3 (EG 15% + LH 7.5%), and KP4 (EG 15% + LH 15%). Observations on the density, follicular structure and protein expression of Bax, Bcl2 and Caspase3 were carried out on ovarian sections made by paraffin method with Hematoxylin-Eosin (HE) staining and immunohistochemistry. The primary antibodies used were Rabbit Anti-Bax polyclonal antibody (A00183 Boster, USA), Rabbit Anti-Bcl-2 (A00040-2 Boster, USA) and Active Caspase-3 Rabbit Polyclonal Antibody (ab4051 Abcam, UK) and One Step Neopoly Detection System Kit (BGNK-0025 Biogear, USA). Identification of each type of preantral follicle using a light microscope connected to Image Raster software and an IHC profiler. The results showed that the cryoprotective effect of longan honey could increase follicle density, G2 and G3 intact follicle index, decrease G1 follicle index, suppress Bax protein expression and caspase 3 and increase Bcl2 protein expression. Thus, longan honey has the potential to be developed as a natural extracellular cryoprotectant in ovarian vitrification applications."

"Radiasi merupakan terapi pilihan untuk kanker serviks stadium III B, namun permasalahan timbul karena adanya sifat radioresisten. Sel punca kanker SPK merupakan salah satu faktor yang diduga berkontribusi terhadap hal tersebut. SOX2 dan OCT4 merupakan faktor transkripsi yang mengekspresikan sifat-sifat SPK, yaitu mengontrol sifat pluripoten, self-renewal, berperan pada karsinogenesis, metastasis, resistensi terhadap terapi dan rekurensi tumor. Faktor apoptosis, DNA repair dan telomerase merupakan mekanisme yang berkaitan dengan radioresisten. Penelitian ini bertujuan untuk mempelajari hubungan antara SOX2 dan OCT4 sebagai penanda SPK terhadap respons terapi radiasi, serta kaitannya dengan faktor apoptosis caspase-3 , DNA repair Chk1 dan telomerase hTERT .Penelitian ini merupakan case control, terhadap 48 kasus karsinoma sel skuamosa serviks stadium III B yang telah menjalani terapi radiasi/kemoradiasi di RS Cipto Mangunkusumo/FKUI. Kasus dibagi dalam 2 kelompok, yaitu hasil terapi komplet 27 kasus dan hasil terapi inkomplet 21 kasus . Kasus dengan respons awal terapi radiasi baik dilakukan pemeriksaan bulan Pap smear dan HPV pada bulan ke-6 atau sampai ke-12 setelah terapi. Ekspresi SOX2, OCT4, caspase-3, Chk1 dan hTERT diperiksa secara imunohistokimia dari blok parafin biopsi awal.Ekspresi kuat SOX2 dan OCT4 dengan H-score masing-masing lebih dari 96,6 dan 61,9 mempunyai hubungan bermakna dengan respons awal terapi radiasi maupun respons akhir terapi radiasi SOX2 p = 0,017, p = 0,004 dan OCT4 p < 0,001, p < 0,001 . Ditemukan hubungan bermakna antara ekspresi Chk1 dan hTERT dengan respons awal terapi radiasi Chk1 p = 0,006, hTERT p = 0,029 . Tidak ditemukan hubungan yang bermakna antara ekspresi caspase-3, Chk1, hTERT dengan ekspresi SOX2 dan OCT4. Uji multivariat menunjukkan bahwa SOX2 dan OCT4 yang paling memengaruhi respons terapi OR = 5,12, p = 0,040 dan OR = 17,03, p < 0,001, secara berurutan . Uji probabilitas menunjukkan kemungkinan respons akhir terapi radiasi inkomplet sebesar 87,91 bila ekspresi kedua penanda SPK kuat.Ekspresi kuat SOX2 dan OCT4 dapat memprediksi hasil terapi radiasi inkomplet pada karsinoma serviks stadium III B.

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Radiotherapy is the main choice of treatment for stage III B cervical cancer, but radioresistance becomes a difficult matter. Cancer stem cell is one of the factors suspected involving in radioresistant cancers. SOX2 and OCT4 are transcription factors which have pluripotent cell characteristics, and self renewal ability. They also involved in carcinogenesis, metastasis, tumor recurrent, and resistance toward therapy. Apoptotic, DNA repair, and telomerase factors are mechanisms that also contribute to radioresistance. This study aims to know the role of SOX2 and OCT4 as CSC markers, apoptotic factor caspase 3 , DNA repair Chk1 and telomerase hTERT toward radiotherapy.The design of this study was case control with 48 cases of stage III B cervical squamous cell carcinoma patients who had finished receiving radiation chemo radiation therapy at Cipto Mangunkusumo Hospital FMUI, Jakarta. They were classified in 2 groups based on the final response of treatment, which were complete and incomplete one. Pap smear and DNA HPV were performed in month 6 or until month 12 after therapy for good initial therapy. Immunohistochemistry was done to analyze SOX2, OCT4, caspase 3, Chk1 and hTERT expression from the paraffin block of initial biopsy.Strong expression of SOX2 and OCT4 with each H score was higher than 96.6, and 61.9 had significant association with both initial and final therapy response SOX2 p 0.017, p 0.004 and OCT4 p 0.001, p 0.001, repectively . There was significant association between expression of Chk1 and hTERT, and initial therapy response p 0.006 for Chk1, and p 0.029 for hTERT . No significant differences were found between caspase 3, Chk1, hTERT, and SOX2 and OCT4. Multivariate analysis showed SOX2 and OCT4 were the most influenced antibodies for radiotherapy response OR 5.12, p 0.040, and OR 17.03, p 0.001, respectively . The likelihood of incomplete final therapy response was 87.91 if the expression both of CSC markers were strong.Expression of SOX2, and OCT4 could predict the incomplete radiotherapy of stage III B cervical cancer cases. "

"Latar Belakang: Pencucian spermatozoa dengan metode Swim-Up SU dan Density Gradient Centrifugation DGC untuk menyeleksi spermatozoa motil telah lama dilakukan, akan tetapi angka keberhasilan masih tergolong rendah. Alpha lipoic acid ALA merupakan antioksidan biologis poten yang berperan dalam regulasi serangan radikal bebas dan pencegahan terhadap kerusakan oksidatif yang disebabkan oleh peroksidasi lipid yang dapat berkontribusi pada integritas DNA spermatozoa. Selain itu, prolaktin PRL adalah salah satu hormon peptida yang juga merupakan faktor prosurvival spermatozoa melalui mekanisme supresi terhadap aktivasi protein kaspase, sehingga berperan dalam proteksi terhadap integritas DNA spermatozoa. Penelitian ini bertujuan untuk mengevaluasi pengaruh pemberian ALA dan PRL terhadap indeks fragmentasi DNA IFD dan status apoptosis spermatozoa deteksi protein kaspase setelah dilakukan pencucian spermatozoa dengan metode SU dan DGC. Metode: Sampel semen diperoleh dari 23 pria normozoospermia dari pasangan wanita infertil dan menjalani IUI. Analisis semen terhadap motilitas dan kecepatan dilakukan sebelum dan sesudah pencucian. Setelah pencucian spermatozoa dengan SU dan DGC, sampel kemudian diinkubasi pada berbagai konsentrasi ALA yaitu 0,625 mg ALA 1 , 1,25mg ALA 2 , 2,5 mg ALA 3 dan pada berbagai konsentrasi PRL yaitu 500 ng PRL 1 , 750 ng PRL 2 , serta 1000 ng PRL 3 . Selanjutnya dilakukan uji sperm chromatin dispersion SCD untuk mengevaluasi fragmentasi DNA spermatozoa dan uji Western Blot untuk mendeteksiprotein kaspase. Hasil: Studi menunjukkan bahwa tingkat IFD spermatozoa setelah pemberian ALA dan PRL mengalami penurunan dibandingkan dengan sampel semen setelah dilakukan pencucian, bahkan dibandingkan dengan sampel semen sebelum pencucian.Protein kaspase ditemukan pada sampel semen sebelum pencucian maupun setelah pencucian dengan metode SU dan DGC. Metode SU dapat menyeleksi spermatozoa dengan IFD yang lebih rendah dibandingkan metode DGC pada konsentrasi optimal ALA dan PRL. Kesimpulan: ALA dan PRL terbukti dapat menyeleksi spermatozoa dengan kualitas spermatozoa yang lebih baik ditinjau dari indeks fragmentasi DNA dan level apoptosis yang lebih rendah, setelah dilakukan pencucian.

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Background: Several methods were done to improved the success rate of intra uterine insemination IUI , some of them are Swim Up SU and Density Gradient Centrifugation DGC sperm preparation, nevertheless the success rate still remain low. Alpha Lipoic Acid ALA is a potent biological antioxidant that play role in regulation of free radical attack and oxidative damage prevention caused by lipid peroxidation which ultimately contribute to DNA integrity of the sperm. Moreover, prolactin PRL is one of peptide hormones which also a prosurvival factor of sperm through suppression of activation of caspase protein mechanism, thus affecting sperm DNA integrity. This study aimed to evaluate the effect of ALA and PRL supplementation on DNA fragmentation DFI and apoptotic stateof the spermafter sperm preparation using SU and DGC methods.

Methods: Semen samples were obtained from 23 men normozoospermia from partners of women who infertile and underwent IUI. Semen analysis was performed for motility and velocity before and after sperm preparation. After SU and DGC sperm preparation, samples were incubated in concentration of ALA at 0,625 mg ALA 1 , 1,25mg ALA 2 ,2,5 mg ALA 3 and PRL at 500 ng PRL 1 , 750 ng PRL 2 , 1000 ng PRL 3 . The Sperm Chromatin Dispersion SCD test was performed to evaluate the sperm DNA fragmentation and Western Blot assay to detect the caspase protein.

Results: This study confirmed that the level of sperm DNA fragmentation index DFI of sperm after supplementation of ALA and PRL were decreased compared to the sperm after preparation even compared to the whole semen. The presence of caspase protein was detected in whole semen samples,and after sperm preparation both SU and DGC, yet SU method could select the sperm with lower level of DNA fragmentation than the DGC method, both at the optimum concentration of ALA and PRL.

Conclusions: ALA and PRL were proved to select the better sperm quality with lower level of sperm DNA fragmentation and minimum density of caspase protein after sperm preparation."