Investigation of the protective effects of citicoline on peripheral nerve injuries

CDP-kolin (sitikolin), fosfolipid metabolizmasında yer alan önemli bir endojen metabolittir. Nöral iyileşmede, iskemik nöral hastalıklarda, nörodejeneratif hastalıklarda, nöropatik ağrıda, bazı kardiyovasküler ve endokrin hastalıklarda yararları bilinmektedir. MMP ve TIMP, hücre dışı matriksin remodelizasyonunda anahtar role sahip proteinlerdir. Bu çalışmada CDP-kolinin, siyatik sinirin transseksiyon-sütürasyon modeliyle oluşturulan nöral hasar sonrası bu moleküller üzerindeki etkisi araştırıldı. Çalışmada 108 adet Wistar albino erkek sıçan kullanıldı. Hayvanlar eşit sayıda üç gruba ayrıldı. Birinci grupta sağ siyatik sinir cerrahi olarak ortaya kondu ancak transseksiyon ve enjeksiyon yapılmadı (Sham grubu). İkinci ve üçüncü gruplarda sağ siyatik sinire transseksiyon ve sütürasyon uygulandı. İkinci grupta 3 ml salin, üçüncü grupta 0,3 ml CDP-kolin işlem gününde intraperitoneal olarak enjekte edildi. Her grup üçer alt gruba ayrılarak işlemin 1. 3. ve 7. günlerinde sağ siyatik sinirler eksize edildi. Hayvanların yarısına eksizyondan önce anatomik çalışma amacıyla perfüzyon işlemi yapıldı. Alınan siyatik sinir örneklerinde Western blot yöntemi ile aktin, MMP-9, MMP-2, TIMP-1 ve TIMP-3 protein konsantrasyonlarına ve Zimografi yöntemiyle MMP-9 ve MMP-2 aktivitelerine bakıldı. Aynı zamanda anatomik olarak akson iyileşmesi incelendi. MMP-9 ve MMP-2 miktarları ve aktiviteleri siyatik sinir hasarını takip eden 1. günde Salin ve CDP-kolin gruplarında Sham grubuna göre anlamlı olarak artarken, 3. ve 7. günlerde CDP-kolin grubunda Salin grubuna göre anlamlı olarak azaldı (p<0.001). TIMP-1 ve TIMP-3 miktarları ise siyatik sinir hasarını takip eden 1. günde her üç grupta da değişmezken, 3. ve 7. günlerde CDP-kolin grubunda Sham ve Salin gruplarına kıyasla anlamlı olarak artış gösterdi; Salin grubunda ise Sham grubuna göre hiçbir dönemde anlamlı değişiklik olmadı. Histomorfolojik değerlendirmede, CDP-kolin grubunda yeni akson oluşumları, adacık formasyonları (kompartmanlar) şeklinde akson toplulukları ve bu aksonlar üzerinde myelin oluşumunun başladığı ve ilerlediği gözlemlendi. Myelinize akson dansitesinin Salin grubunda zamanla kötüleşerek azaldığı, sham grubunda anlamlı bir değişim göstermediği tespit edildi. CDP-Kolin grubunda ise her üç zaman diliminde alınan örnekler salin grubu ile karşılaştırıldığında anlamlı derecede iyileşme ve artmış myelinli akson dansitesi gösterdiği görüldü. Çalışmamızda sıçan siyatik sinirinin primer kesi ve sütur modelinde CDP-kolin tedavisinin MMP-9 ve MMP-2 düzeylerini ve aktivitelerini azalttığı ve TIMP-1 ile TIMP-3 düzeylerini arttırdığı literatürde ilk kez gösterilmiştir. Çalışmamız periferik sinir kesilerinde CDP-kolin uygulaması ile sağlanan nöroproteksiyonun etki mekanizmasının aydınlatılması bakımından önemlidir. Bulgularımız gelecekte CDP-kolin'in periferik sinir hasarındaki etkinliğinin inceleneceği klinik çalışmalara ışık tutar niteliktedir.CDP-choline (citicoline) is an endogenous metabolite which plays an important role in phospholipid metabolism. It has known beneficial effects in neural trauma, ischemic conditions such as stroke and cerebral infarcts, neurodegenerative conditions such as Alzheimer's or Parkinson's disease, and neuropathic pain, as well as various cardiovascular and endocrine diseases. MMPs and TIMPs are key molecules in the remodelling of the extracellular matrix. In the current study, the effect of CDP-choline on these molecules after sciatic nerve injury in the transsection-suturation model in rats was evaluated. 108 Wistar albino male rats were included in the study. Rats were divided into three groups of equal quantities. The first group was composed of rats whose right sciatic nerves were surgically exposed but no transsection or injection was made (the Sham group). The rats in the second and third groups underwent transsection and immediate primary suturation of the right sciatic nerve. 3 ml of normal saline and 0.3 ml of CDP-choline were injected intraperitoneally to rats in the second (the Saline group) and the third (the CDP-choline group) groups, respectively, at the sime time with the intervention. Each group was further divided into 3 subgroups, of which the right sciatic nerves were excised at the 1st, 3rd and 7th days after the initial intervention. Half of the animals underwent perfusion process for the purpose of anatomic study. The sciatic nerve specimens were then analyzed with the Western blot technique for levels of MMP-9, MMP-2, TIMP-1, TIMP-3, and actin and with Zymography method for MMP-9 and MMP-2 activities. Axonal regeneration was also studied anatomically. Levels and activities of MMP-9 ve MMP-2 were significantly higher in the Saline and CDP-choline groups than in the Sham group on the 1st day postinjury, albeit being significantly higher in the CDP-choline group than in the Saline group on the 3rd and 7th days postinjury (p<0.001). On the other hand, levels of TIMP-1 and TIMP-3 were not different among the three groups on the 1st day postinjury, but they were significantly higher in the CDP-choline group than that of the Sham and Saline groups. No significant difference was observed between the Sham and the Saline groups throughout the study period (i.e. 1st or 3rd or 7th days). On histomorphological evaluation, new axon formation, and formation and advancement of myelination on newly formed islets (compartments) of axonal regrowth were observed in the CDP-choline group. Myelinated axon density was increasingly deteriorating in the saline group while it was steady and significantly increasing in the saline and the CDP-choline groups respectively. In our study, for the first time in the literature, CDP-choline was shown to decrease MMP-2 and MMP-9 levels and activities while increasing TIMP-1 and TIMP-3 levels in rat sciatic nerve transection and suturation model. These data are significant in terms of revealing the mechanism of action of CDP-choline in providing neuroprotection in peripheral nerve transection. Our findings may shed light on future clinical studies investigating the effect of CDP-choline in peripheral nerve injuries

Intracranial Pressure Measurement of and Investigation of the Effect of Aquaporin on Hydrocephalus Induced by Experimental Kaolin and Autologous Blood Injection: An Animal Study Deneysel Kaolin ve Otolog Kan Enjeksiyonu ile İndüklenen Hidrosefali Üzerine Aquaporin Etkisinin İntrakraniyal Basınç Ölçümü ve Araştırılması: Hayvan Deneyi

Objective: Hydrocephalus is a condition in which brain tissue is damaged due to ventricular enlargement. In experimental mod-els, hydrocephalus was induced by injecting various substances into the cerebrospinal fluid pathway or creating a subarachnoid hemorrhage model. Material and Methods: Five experimental groups were formed. The stereotaxic frame was placed in accordance with the coordinates calculated for the cisterna magna. In Group 1, only a spinal puncture was performed. In Group 2, a hydrocephalus model was created by injecting kaolin (Group 2A) and autologous blood (Group 2B). A hydrocephalus model was created with kaolin in Group 3, autologous blood in Group 4, and acetazolamide treatment was applied to both groups post-injection. Autologous blood was taken from the experimental groups before de-capitation, and the levels of tumor necrosis factor-alpha (TNF-α) and interleukin (IL)-1 were measured by the ELISA method. After histological staining, the lateral ventricle size was measured. Intracranial pressure (ICP) measurements were taken on days 0 and 7 in all groups. Results: There was a significant increase in ICP in Groups 2A and 2B. TNF-α and IL-1 values increased more significantly in the groups that did not receive acetazolamide treatment compared to the group that received treatment. Conclusion: There was an increase in ventricle dimensions and ICP as well as TNF-α and IL-1 levels in both hydrocephalus models. Ac-etazolamide treatment was seen to be significantly more effective in kaolin group. This study is important because it is the first in the literature to perform biochemical and histopathological examination and ICP measurements all in the same hydrocephalus model