全國中小學科展

醫學與健康科學

新式高能磁場對於骨髓幹細胞軟骨分化的影響

近年來,再生醫療利用幹細胞幫助組織器官新生的方法已有長足進步,但仍然面臨許多的瓶頸。例如: 幹細胞注射進入人體以後,無法繼續分化生長成所需組織。最近,高能磁場的使用也是醫學界蓬勃發展的議題,我們試圖利用高能磁場誘導幹細胞的分化,希望可以增加幹細胞分化的效能,這是一創新的嘗試。我們比較新型高能磁場(RMS)與對照組於骨髓幹細胞(BMSCs)的分化效果,發現RMS 中叢集式刺激(iTBS)的SOX9和COL10的基因表現與免疫組織化學染色明顯優於15Hz RMS 與對照組。最佳的刺激設計為強度50%的iTBS連續施打五天。我們亦發現每回合900 pulses有優於每回合600 pulses的趨勢。此實驗證實iTBS能夠增加BMSCs在體外的軟骨分化表現,反映在基因表現層次到膠原蛋白合成的層次。未來,此方法將進一步使用於動物實驗,期望擴展到臨床上用於退化性關節炎的治療,對再生醫療發展有所幫助。

Development of a neurointerface glove with tactile feedback

Research Question or Engineering Problem A stroke continues to be the most important medical and social problem of the modern world. Stroke is a type of acute cerebrovascular accident (ACVA) and is characterised by a sudden (within minutes, less often - hours) appearance of focal neurological symptoms (motor, speech, sensory, coordinating, visual and other disorders) and / or general brain disorders (depression of consciousness, headache, vomiting, etc.) that persist for more than 24 hours or lead to death of the patient in a short period of time due to a cause of cerebrovascular origin. There are two clinical and pathogenetic forms of stroke: ishemic stroke (cerebral infarction) is caused by acute focal cerebral ischemia, leading to infarction (zone of ischemic necrosis) of the brain; hemorrhagic stroke (non-traumatic intracerebral hemorrhage) is caused by rupture of an intracerebral vessel and blood penetration into the brain parenchyma or rupture of an arterial aneurysm with subarachnoid hemorrhage (SAH). ACVA also includes transient disorders of cerebral circulation, characterised by the sudden occurrence of focal neurological symptoms that develop in a patient with cardiovascular disease (arterial hypertension, atherosclerosis, atrial fibrillation, vasculitis, etc.), last for several minutes, less often hours, but no more than 24 hours, and ends with a full restoration of the impaired brain functions. Transient disorders of cerebral circulation include: transient ischemic attack (TIA), which develops as a result of short-term local cerebral ischemia and is characterised by sudden transient neurological disorders with focal symptoms; hypertensive cerebral emergency, which is a condition associated with an acute (usually significant) rise in blood pressure (BP) and accompanied by the appearance of general cerebral (less often focal) neurological symptoms secondary to hypertension. The most severe form of hypertensive crisis is acute hypertensive encephalopathy, the basis of pathogenesis of which is cerebral edema. Cerebral infarction generally is the result of the interaction of many etiopathogenetic factors, which can be subdivided into local and systemic ones. Local factors include: morphological changes in the brachiocephalic or intracerebral arteries (pathological tortuosity, etc.), atherosclerotic lesions of the vessels of the aortic arch and cerebral arteries, cardiac lesions as a source of thromboembolic cerebral infarctions, fibromuscular dysplasias of the walls of the brachiocephalic and cerebral arteries, brachiocephalic artery dissection, vasculitis (arteritis), changes in the cervical spine with the formation of extravasal compression of the vertebral arteries, anomalies in the structure of the vessels of the neck and brain (hypoplasia of the vertebral artery, trifurcation of the internal carotid artery), etc. Systemic factors include: disorders of central and cerebral hemodynamics (a sharp change in BP or a decrease in cardiac output, etc.), hereditary and acquired coagulopathies, polycythemia, certain forms of leukemia, hypovolemia, psychoemotional stress / distress, etc., hypercoagulative / hyperaggregatory side effects of a number of medications (oral contraceptives, etc.). In the 俄羅斯n Federation, more than 500 thousand people have a stroke every year. About 25,000 new cases of stroke occur in St. Petersburg every year. The incidence of stroke in the 俄羅斯n Federation is 3.48 ± 0.21 cases per 1000 people. The incidence of various types of ACVA varies widely, in particular, cerebral infarctions account for 65–75%, hemorrhages (including subarachnoid hemorrhages) – 15–20%, transient cerebral circulation disorders account for 10–15%. The frequency of cerebral strokes in the population over 50–55 years old increases by 1.8–2 times in each subsequent decade of life. Medical and socio-economic consequences of ACVA are very significant, in particular, death in the acute period of stroke occurs in 34.6% cases, during the first year after the end of the acute period in 13.4% cases; severe disability with the need for constant care is present in 20.0% of stroke patients; 56.0% have limited working capacity and only 8.0% return to their previous work activity. Disability due to stroke (the national average is 56–81%) in our country ranks first among all causes of primary disability, amounting to 3.2 per 10 thousand people. Stroke mortality among working-age population has increased in the 俄羅斯n Federation by more than 30% over the past 10 years. The annual death rate from stroke in our country is 175 per 100 thousand people. Stroke annually becomes the main cause of disability: 85% of victims experience a decrease in strength or a complete lack of ability to control the muscles of half of the body and only half of them recover limb functions partially or completely; the rest of those who have suffered a stroke remain paralysed and require care, since they are not able to completely independent existence. In this regard, recently, in the process of rehabilitation, the technology of brain-computer interfaces (BCI) has begun to be actively used. on the basis of this technology exercise machines are created. These exercise machines are controlled directly by the patient himself. This feature of the technology increases the effect of the procedure by providing a direct connection between the patient's desire and effort with the work of the simulator. The greatest development of this technology is observed in the field of medicine, where BCIs are used as a means of communication or as one of the tools of neurorehabilitation. In this regard, it seems very promising to develop the most optimal brain-computer interfaces. The goal of our project was to create an automated training complex in the form of a neuro-controlled glove with tactile feedback, designed to simplify access to rehabilitation means.

探討雙黃酮類CRYT對抑制發炎反應與免疫細胞移動的影響

發炎反應與癌症具有密切的關係,而如何抗發炎是現今重要的研究主題。本研究主要探討雙黃酮類藥物CRYT對LPS誘發之發炎反應與免疫細胞遷移的影響。本研究首先以細胞試驗分析細胞激素與趨化因子的表現量觀察CRYT對發炎反應的影響。接著,利用反轉錄聚合酶連鎖反應 (RT-PCR) 和共軛焦顯微鏡觀察黏附分子與調控細胞骨架分子的表現量,以了解CRYT對免疫細胞遷移的影響。再來我們以西方墨點法分析免疫反應調控路徑之蛋白質表現量來研究CRYT的作用途徑。同時,透過程式模擬CRYT的標的蛋白,並用細胞轉染與雙冷光技術驗證模擬結果。本研究發現,CRYT會抑制TNF-、IL-1及IL-6三種促發炎因子與CXCL1、CCL3及CCL4三種趨化因子的表現量。另外,CRYT會抑制黏附分子LFA-1與VLA-4與調控細胞骨架分子Cdc42、PAK1、WASP、Rac1及LIMK1之mRNA表現量,以阻礙F-actin進行聚合作用。本研究也發現,CRYT能藉由抑制NF-B的表現量,進而影響巨噬細胞的發炎作用與細胞移動。在作用機制的方面,CRYT會結合在雌激素受體的配體結合位,並抑制雌激素受體的轉錄作用,而抑制發炎作用與細胞轉移。結論得知,CRYT不但可以抑制發炎反應,還能有效抑制免疫細胞的遷移。

探討吸菸調控絲胺酸合成路徑影響肺癌生長及抗藥性

肺癌為全球死亡率最高的癌別之一。抽菸,肺癌的主要危險因子,臨床上造成抗癌藥效不佳並導致病患的低存活率與不良預後。然而,抽菸影響肺癌的機制仍不清楚。代謝重整最近被視為是癌症的新興特點。絲胺酸合成路徑為葡萄糖代謝的分支之一,參與生物合成材料之製造,並和癌症的惡化有密切的關連性,但缺乏詳細的相關研究。本研究探討抽菸是否透過影響絲胺酸合成路徑來導致肺癌生長,並測試絲胺酸合成路徑抑制劑是否能增強化療藥物吸菸相關肺癌細胞的治療效果。我們的研究發現,抽菸和絲胺酸合成路徑在肺癌中有正相關性且與臨床上的低存活率有關,並證實抽菸調控絲胺酸合成路徑而促進肺癌生長與化療抗性,此現象可因合併給予絲胺酸合成路徑抑制劑而獲得緩解。此研究成果顯示抑制絲胺酸合成路徑可能成為治療吸菸相關肺癌的新策略。

PP5-AMPK Pathway mediates Palbociclib-induced Cell Death in Lung Cancer

肺癌是目前全世界首要致死的癌症。雖然現有的標靶治療與免疫療法已經改善某些病患的情況,但有許多病患仍無有效治療方法。因此,我們急需探求新的藥物治療方法來改善肺癌治療的成效。 在此我們選擇一個新標靶藥物Palbociclib來進行研究,Palbociclib 是剛核准的乳癌藥物,主要是藉由抑制CDK4/6來延緩乳癌生長。我們發現Palbociclib能有效地抑制肺癌細胞株的生長和死亡。特別的是,Palbociclib不僅誘發肺癌細胞凋亡(apoptosis),亦誘發肺癌細胞自噬死亡(Autophagy)。因為AMPK能同時影響細胞凋亡與自噬死亡,我們進一步研究AMPK在Palbociclib誘發癌細胞死亡中的角色,發現Palbociclib主要是藉由抑制去磷酸酶PP5的活性,來增加AMPK的磷酸化,進而活化AMPK,誘發肺癌細胞產生凋亡與自噬。這些藥效原非抑制CDK4/6會產生的作用,我們發現Palbociclib藉由調控PP5-AMPK路徑以誘發細胞死亡。希望這些研究結果能幫助肺癌的臨床研究,以造福病患。

探討Naa10p突變對神經發育的影響

Naa10p (N-alpha-acetyltransferase 10 protein) 突變患者多半有智力障礙的問題,如:奧格登症候群患者常有早夭、腦萎縮及其他發育問題。Naa10p 是一種 N-α-乙醯轉移酶,負責新生肽鏈 N-α-乙醯化。人體中約有 80% 的蛋白質經乙醯化修飾,其中 46% 為 NatA 複合體執行,而 Naa10p 即是 NatA中執行乙醯化的蛋白酶。先前研究顯示Naa10p會影響神經發育,但對Naa10p影響神經發育的機制並不清楚。本研究利用CRISPR/Cas9製作Naa10p的小鼠胚胎幹細胞(mESC)突變株,觀察突變株細胞分裂速率與分化型態的差異,並分析胚胎發育過程Naa10p以及相關基因的表現。 本研究結果證明特定Naa10p突變會加快mESC生長,使其分化為神經母細胞的異常。而其中V111G突變株的Naa10p無法和HYPK結合,且突變S37P及R116W使Naa10p與Naa15p的結合降低,推論此結果可能造成NatA無法正常形成,並改變其與核糖體蛋白的親和力,影響蛋白質乙醯化。目前我們發現突變株中,與細胞分化相關的基因:Oct4、Pax6皆表現異常。以上結果可能可以初步解釋Naa10p突變造成人類疾病的機制。

THE DESIGN OF MICROFLUIDIC PUMP (MFP) FOR MEDICAL FIELD

The ability of microfluidic (MF) device technologies to provide a lot of information with a small amount of sample, the opportunities it offers increases their use in the medical field in the bedside monitoring in drug delivery systems. Three-dimensional (3D) printer technologies provide advantages such as cost-effectiveness in the production of MF devices and quick and easy production in intricate designs. In our project, it is aimed to design microfluidic pumps (MFP) to be used in the medical field and conduct its production with 3D printer technologies. The developed MFP is intended to be at low cost, bio-compatible, adaptable, and portable to the drug, suitable flow properties as a pharmaceutical pump. First of all, MFP air channel, flow channel, etc. parts were designed and printed with the help of a 3D printer and on AutoCAD, one of the professional drawing programs. The poly(dimethylsiloxane) (PDMS) membrane that will enable MFP activation is produced in different thicknesses and glued to the air channel of MFP. The resistance to the applied pressure is observed, and the appropriate membrane thickness is determined as ~ 235µm. Liquid PDMS was applied to the inner surfaces of MFP's air and flow channel, PDMS membrane was placed between them, and the parts were assembled in the oven at 60ºC. MFP has been connected to the pneumatic valve system, where operation codes have been prepared with Arduino Uno, and flow properties have been examined. The flow rate of MFP is ~ 50 µL/min at a maximum of 15 Hz, and the backpressure is ~ 0.085 Pa under a maximum pressure of 3 bar. Also, values such as size, membrane thickness, and applied pressure for the possible models of MFP were supported by theoretical calculations. As a result, MFP, which is biocompatible, drug adaptable, portable, wearable technology application potential, and has suitable flow characteristics as a pharmaceutical pump, has been developed. MFP introduced a microfluidic pump system that can make life easier for the patient and contribute to the national economy through domestic production and can be used as a drug pump in the treatment of diseases such as diabetes and cancer.

Consensus-based Machine Learning Model in the Scoring for Hepatic Steatosis Severity

研究目的:脂肪肝的盛行率在全世界的現代化國家都在增加。病理醫師在評估脂肪肝的嚴重等級時,常缺乏參考意見以減少差異。許多地區也缺乏病理醫師。本研究旨在建立一個有效評估脂肪肝嚴重程度的病理組織學人工智慧模型。研究過程:本研究經由臺中榮民總醫院人體研究倫理審查委員會審核通過,篩選後取得病理資料庫中適合的肝臟組織切片,由兩位病理醫師獨立為肝組織中脂肪堆積的程度評分。再以病理醫師討論後的共識答案為分級標準,來訓練人工智慧模型。研究結果:在100個樣本的獨立測試中,人工智慧模型和病理專科醫師的評分,有71%完全相同、27%差異只有一個等級、2%差異2個等級、而沒有2個等級以上的差異。結論及應用:我們已初步建立一個可以評估脂肪肝嚴重程度的人工智慧模型。這模型可為將來人工智慧的臨床應用,建立一個良好的基礎。

Anti-bacterial Crab bio-bandages with Bio-dressings 2.0

Commercially available bandages such as hydrocolloid are neither biodegradable nor anti-bacterial. Chitin is known to be the second most naturally available polysaccharide which could be transformed to chitosan which is known to be anti-bacterial (Hasan, 2018) (Chao, 2019) and haemostatic (Okamoto, 2003) (Hu, 2018). Chitosan can be further converted to hydrogel which is bio-degradable and has good water absorbance. Anti-bacterial crab bio-bandages and crab bio-dressings should be bio-degradable as it took 42 days and a month for complete bio-degradation respectively, so they should be better than commercial bandages such as Nexcare Hydrocolloid as the disposal of anti-bacterial crab bio-bandages with bio-dressings would no longer pose burden to landfilling or threat to our environment. Anti-bacterial crab bio-bandages with bio-dressings are anti-bacterial with degree of deacetylation of DD% (measured using FTIR Spectrum II) 82.6% (due to the presence of chitosan) even without the application of other anti-bacterial agents and hence can provide complete protection of wounds from skin and soft tissues infections and haemostatic (due to the presence of chitosan). After testing and certification based on IS997:2004 and BS EN 13726-1, they should meet many requirements specified. Anti-bacterial crab bio-bandages should be eligible for marketing. Some results were as follows: 1.4 Anti-bacterial effect of crab hydrogels and roasted crab hydrogels Pure chitosan, crab chitosan, crab hydrogels and roasted crab hydrogels showed significant anti-bacterial effect. NO oral bacterial colonies were present in drinking water with crab hydrogels. Thus crab hydrogels could serve as effective anti-bacterial wound dressings. 1.6 Basing on IS997:2004 standard, the load per unit of area of anti-bacterial bio-bandages was 342g/m2 which met the minimum requirement of 36g/m2, the anti-bacterial bio-bandages had stronger tension strength (>20N both in dry and wet conditions) than commercial hydrocolloid. (2.7N dry 2.8N wet) which was comparable with that required (50-67N) and pH of about 7 which met the pH range of 4.5-8. 1.7 The FSA Free-Swell Absorbency of synthetic blood of crab hydrogel bio-dressings was 1.86g per 5cm x 5cm dressing which was much higher than that of commercial hydrocolloid (0.299g per 5cm x 5cm dressing) based on BS EN 13726-1.

HOST TARGET PROTEINS OF SPIKE PROTEIN OF SARS-COV-2

Coronavirus Disease 2019 (COVID-19) is a newly emerged infectious disease caused by the new severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV-2). In less than one year, the virus has spread around the entire world, killing millions of people and disrupting travel and business worldwide. During infection, the virus uses its Spike protein to dock onto the Ace2 protein on the surface of its human host cell. Spike is 1273 amino acids long and only a short fragment of Spike (319-541) is sufficient to bind Ace2. We hypothesized that the remaining protein sequences of Spike might have functions for viral replication beyond the binding of Ace2. We have performed Split-Ubiquitin protein-protein interaction screens to isolate human proteins by their ability to bind to Spike, and we have identified Annexin2A2 and Cytochrome b as novel human protein interaction partners of Spike. Annexin2A2 is involved in both endocytosis and exocytosis, and the protein interaction with Spike might help the virus to enter and exit its host cell. The presence of the mitochondrial Cytochrome b protein inside the cytosol promotes apoptosis, and the protein interaction with Spike could speed up sapoptosis of the infected human cell. The Nub cDNA libraries that we have generated also allowed us to screen for synthetic peptides that interact with Spike. We have isolated two synthetic peptides, FL1a and FL7a, derived from the non-coding parts of human mRNAs by their ability to interact with Spike. We found that both FL1a and FL7a interact with the C-terminal half of the Spike protein. We also found that FL7a is able to block the Spike-Spike self-interaction at the C-terminal half of the Spike protein and we think that this could block the reassembly of the Spike protein in the host cell during viral reassembly. We hope that those synthetic peptides could be used as drugs due to their ability to block protein-protein interactions of Spike with human host proteins that are essential for viral replication.