Recent advances in the diagnostic methods and therapeutic strategies of transthyretin cardiac amyloidosis

Cardiac amyloidosis is a progressive disease characterized by the buildup of amyloid fibrils in the extracellular space of the heart.It is divided in 2 main types,immunoglobulin light chain amyloidosis and transthyretin amyloidosis(ATTR),and ATTR amyloidosis is further divided in 2 subtypes,non-hereditary wild type ATTR and hereditary mutant variant amyloidosis.Incidence and prevalence of ATTR cardiac amyloidosis is increasing over the last years due to the improvements in diagnostic methods.Survival rates are improving due to the development of novel therapeutic strategies.Tafamidis is the only disease-modifying approved therapy in ATTR amyloidosis so far.However,the most recent advances in medical therapies have added more options with the potential to become part of the therapeutic armamentarium of the disease.Agents including acoramidis,eplontersen,vutrisiran,patisiran and anti-monoclonal antibody NI006 are being investigated on cardiac function in large,multicenter controlled trials which are expected to be completed within the next 2-3 years,providing promising results in patients with ATTR cardiac amyloidosis.However,further and ongoing research is required in order to improve diagnostic methods that could provide an early diagnosis,as well as survival and quality of life of these patients.

Repression of retinal microvascular endothelial cells by transthyretin under simulated diabetic retinopathy conditions

AIM： To investigate biological effects of transthyretin （TTR） on the development of neovascularization under simulated diabetic retinopathy （DR） condition associated with high glucose and hypoxia. METHODS： Human retinal microvascular endothelial cells （hRECs） were cultured in normal and simulated DR environments with high glucose and hypoxia. The normal serum glucose concentration is approximately 5.5 mmol/L; thus, hyperglycemia was simulated with 25 mmol/L glucose, while hypoxia was induced using 200 μmol/L CoCI. The influence of TTR on hRECs and human retinal pigment epithelial cells （hRPECs） was determined by incubating the cells with 4μmol/L TTR in normal and abnormal media. A co -culture system was then employed to evaluate the effects of hRPECs on hRECs. RESULTS： Decreased hRECs and hRPECs were observed under abnormal conditions, including high- glucose and hypoxic media. In addition, hRECs were significantly inhibited by 4 pmol/L exogenous TTR during hyperglycemic culture. During co-culture, hRPECs inhibited hRECs in both the normal and abnormal environments. CONCLUSION： hREC growth is inhibited by exogenous TTR under simulated DR environments with highglucose and hypoxic, particularly in the medium containing 25 mmol/L glucose, hRPECs, which manufacture TTR in the eye, also represses hRECs in the same environment. TTR is predicted to inhibit the proliferation of hRECs and neovascularization.

Negative effects of transthyretin in high myopic vitreous on diabetic retinopathy

AIM：To analyze the relationship between vitreous transthyretin（TTR） levels,high myopia and diabetic retinopathy（DR）.METHODS：We selected 6722 individuals from the southern Jiangsu Province for diabetes and ophthalmic examinations.The TTR concentration in the vitreous of 50 patients with high myopia and diabetes,50 patients with only DR,and 20 healthy controls were determined by ELISA.Key factors in Tie2 pathway in DR development including vascular endothecial growth factor（VEGF）,Tie2,Angpt1,Angpt2,vascular endothelial growth factor receptor（VEGFR） 1 and VEGFR2 were also detected by ELISA.RESULTS：The prevalence of DR in patients with diabetes and myopia [〈6.00 diopter（D）],diabetes and high myopia（〉6.00 D）,and diabetes without myopia were 11.1%,2.5%,and 60.0%,respectively.The vitreous TTR concentration of patients with diabetes and high myopia was approximately 6.5-and 4.2-times higher than those of patients with DR and healthy controls,respectively（P〈0.05）.Following the vitreous TTR concentration,the levels of VEGF,Tie2,Angpt1,Angpt2,VEGFR1 and VEGFR2 in vitreous of diabetes and high myopia patients,DR patients and healthy controls were detected as dramatically fluctuated.CONCLUSION：The results suggest that TTR can affect the vitreous contents of key factors in Tie2 pathway for neovascularization,and there should be a protective association between abundant TTR levels in the vitreous of highly myopic patients and a decreased risk of DR.

Transthyretin Arg-83 mutation in vitreous amyloidosis

Both of the patients in the report had floaters and progressive vision loss for years. Two cases of familial vitreous amyloidosis occurred in three generations with typical white fibrilar opacities in the vitreous body. Pars plana vitrectomy was performed in the two patients. The vitreous specimens were subjected to histopathological examination. The specimens showed typical microscopic features of amyloidosis with Congo red stain and non-branching fibrils were seen randomly distributed with 5-10nm in diameter on a transmission electron microscope. All of the exons of the transthyretin gene were amplified with DNA isolated from the peripheral blood cells. Bi-directional sequencing of the transthyretin gene revealed a single base-pair substitution, which results in an amino acid substitution at position83, glycine to arginine (transthyretin Arg-83).

Symptomatic Val122del mutated hereditary transthyretin amyloidosis: Need for early diagnosis and prioritization for heart and liver transplantation

Background: Hereditary transthyretin(ATTRv) amyloidosis is an autosomal dominant disease linked to transthyretin gene mutations which cause instability of the transthyretin tetramer. After dissociation and misfolding they reassemble as insoluble fibrils(i.e. amyloid). Apart from the common Val30 Met mutation there is a very heterogeneous group of non-Val30 Met mutations. In some cases, the clinical picture is dominated by a rapidly evolving restrictive and hypertrophic cardiomyopathy. Methods: A case series of four liver recipients with the highly clinically relevant, rare and particularly aggressive Val122 del mutation is presented. Medical and surgical therapeutic options, waiting list policy for ATTRv-amyloidosis, including the need for heart transplantation, and status of heart-liver transplantation are discussed. Results: Three patients needed a staged(1 patient) or simultaneous(2 patients) heart-liver transplant due to rapidly progressing cardiac failure and/or neurologic disability. Domino liver transplantation was impossible in two due to fibrotic hepatic transformation caused by cardiomyopathy. After a follow-up ranging from 3.5 to 9.5 years, cardiac(allograft) function was maintained in all patients, but neuropathy progressed in three patients, one of whom died after 80 months. Conclusions: This is the first report in(liver) transplant literature about the rare Val122 del ATTRv mutation. Due to its aggressiveness, symptomatic patients should be prioritized on the liver and, in cases with cardiomyopathy, heart waiting lists in order to avoid the irreversible neurological and cardiac damage that leads to a rapid lethal outcome.

Wild type transthyretin amyloidosis,a reason not to be forgotten for heart failure of preserved ejection fraction in the elderly

Amyloidosis is a multisystem disease that is characterized by deposition of fibrils in extracellular tissue,which mainly involves the kidney,heart,and autonomic nervous system.Two types of amyloidosis typically infiltrate the heart,including immunoglobulin light-chain(AL)and amyloid transthyretin(ATTR).ATTR is further subdivided into wild-type ATTR and variant ATTR caused by point mutations in the TTR gene.[1]Wild-type ATTR is considered as not uncommon in older patients with heart failure.Recently,a comprehensive set of consensus recommendations for the suspicion and diagnosis of ATTR was published,with particular focus on the combined application of noninvasive methods.[2]We present here a case of wild-type TTR cardiac amyloidosis(ATTRwt-CA),which was diagnosed by noninvasive modalities,and provide an overview of the recommended diagnostic approach of CA.Furthermore,to the best of our knowledge,this is the first Chinese case of ATTRwt-CA reported to date.

Clinical features of retinal amyloid angiopathy with transthyretin Gly83Arg variant

AIM:To report on the clinical features of patients with retinal amyloid angiopathy(RAA)who were identified to be caused by the transthyretin(TTR)Gly83Arg variant.METHODS:Case series of five patients diagnosed with RAA was collected at Affiliated Hospital of Zunyi Medical University from January 2010 to December 2021.The clinical features,therapeutic strategies,and prognoses of all patients were reviewed.RESULTS:Five patients with a mean age of 52.00±7.23y were diagnosed as RAA.These patients were previously diagnosed with hereditary transthyretin amyloidosis caused by the TTR Gly83Arg variant.Vitreous opacity was found in all 10 eyes,and 7 eyes developed RAA 2 to 20y after the onset of hereditary transthyretin amyloidosis.The clinical manifestations were recurrent vitreous hemorrhage in 2 eyes(29%),neovascular glaucoma in 2 eyes(29%),and iris neovascularization in 1 eye(14%).Microangioma lesions were found in all affected eyes that underwent fundus fluorescein angiography(FFA)in this group of cases,and the incidence of the retinal non-perfusion area was 67%.Although no cases of retinal neovascularization were found,the prognosis of visual acuity was not ideal.CONCLUSION:This is the first report of RAA in patients with the TTR Gly83Arg variant.Complications such as RAA and glaucoma will seriously affect the visual prognosis of patients.Thereafter,regular ophthalmic follow-up of patients with hereditary transthyretin amyloidosis is essential.And FFA after vitrectomy is very important,which can help ophthalmologists detect RAA earlier and treat it in time.

Transthyretin—A Key Gene Involved in Regulating Learning and Memory in Brain, and Providing Neuroprotection in Alzheimer Disease via Neuronal Synthesis of Transthyretin Protein

Transthyretin (TTR), a carrier protein present in the liver and choroid plexus of the brain, has been shown to be responsible for binding thyroid hormone thyroxin (T4) and retinol in plasma and cerebrospinal fluid (CSF). TTR aids in sequestering of beta-amyloid peptides Aβ deposition, and protects the brain from trauma, ischemic stroke and Alzheimer disease (AD). Accordingly, hippocampal gene expression of TTR plays a significant role in learning and memory as well as in simulation of spatial memory tasks. TTR via interacting with transcription factor CREB regulates this process and decreased expression leads to memory deficits. By different signaling pathways, like MAPK, AKT, and ERK via Src, TTR provides tropical support through megalin receptor by promoting neurite outgrowth and protecting the neurons from traumatic brain injury. TTR is also responsible for the transient rise in intracellular Ca2+ via NMDA receptor, playing a dominant role under excitotoxic conditions. In this review, we tried to shed light on how TTR is involved in maintaining normal cognitive processes, its role in learning and memory, under memory deficit conditions;by which mechanisms it promotes neurite outgrowth;and how it protects the brain from Alzheimer disease (AD).

Multi-modality imaging in transthyretin amyloid cardiomyopathy

Transthyretin amyloid(TTR)cardiomyopathy is a disease of insidious onset,which is often accompanied by debilitating neurological and/or cardiac complications.The true prevalence is not fully known due to its elusive presentation,being often under-recognized and usually diagnosed only late in its natural history and in older patients.Because of this,effective treatment options are usually precluded by multiple comorbidities and frailty associated with such patients.Therefore,high clinical suspicion with earlier and better detection of this disease is needed.In this review,the novel applications of multimodality imaging in the diagnostic pathway of TTR cardiomyopathy are explored.These include the complimentary roles of transthoracic echocardiography,cardiac magnetic resonance,nuclear scintigraphy and positron emission tomography in quantifying cardiac dysfunction,diagnosis and risk stratification.Recent advances in novel therapeutic options for TTR have further enhanced the importance of a timely and accurate diagnosis of this disease.

Early and aggressive presentation of wild-type transthyretin amyloid cardiomyopathy:A case report

BACKGROUND Wild-type transthyretin amyloidosis(ATTRwt)is the most common form of transthyretin amyloid cardiomyopathy,occurring mostly over age of 60 years(mean age of 80 years).Mean survival without treatment is 3.6 years,making early detection imperative.We report an unusual case of a 58-year-old patient with ATTRwt cardiomyopathy requiring heart transplantation.CASE SUMMARY A 58-year-old male presented with progressive fatigue,shortness of breath,weight gain,leg swelling,orthopnoea,and paroxysmal nocturnal dyspnoea for several months.Approximately ten months before this clinical presentation,the patient had first received a diagnosis of heart failure with reduced ejection fraction(EF)of 15% to 20%.The patient was started on appropriate guidelinedirected medical therapy with only mild improvement in his EF.Upon further investigation,echocardiogram,technetium pyrophosphate scan(Tc PYP),and cardiac magnetic resonance imaging(cMRI)suggested a diagnosis of amyloidosis,and ATTRwt was subsequently confirmed with native heart tissue biopsy,congo red staining,liquid chromatography-tandem mass spectrometry,and genetic testing.The patient was successfully treated with heart transplantation and is doing well post-transplant.CONCLUSION Wild-type ATTR amyloidosis should be kept on differentials in all patients(even less than 60 years old)with non-ischemic cardiomyopathy,especially in the setting of increased ventricular wall thickness and other classic echocardiogram,cMRI,and Tc PYP findings.Early diagnosis and management can be consequential in improving patient outcomes.

Vitreous amyloidosis caused by a Lys55Asn variant in transthyretin: A case report

BACKGROUND Amyloidosis is caused by misfolding of proteins and is characterized by formation of extracellular aggregates of insoluble fibrin.The primary effects in the eye include sharp deterioration of visual acuity as a result of vitreous opacity.According to the local and systemic distribution characteristics of amyloid deposits and their fibrin components,amyloidosis can be classified as primary,secondary or familial.Therefore,we report a typical case of vitreous amyloidosis in hereditary transthyretin amyloidosis(hATTR)to improve ophthalmologists’understanding of the disease and reduce misdiagnosis and recurrence.CASE SUMMARY The patient was a 49-year-old man who complained of progressive visual decline in both eyes over a 2-mo period.No systemic diseases such as diabetes or hypertension were reported,and no obvious family history of disease was identified.The patient’s visual acuity was HM/10 cm in the right eye and 0.06 in the left eye.He had a transparent cornea in both eyes,with a normal anterior depth,clear aqueous humor,no obvious iris abnormalities,round pupils of approximately 3 mm in diameter,normal direct and indirect light reflexes,and normal intraocular pressure.After various examinations,the patient was diagnosed with binocular vitreous amyloidosis secondary to hATTR associated with a Lys55Asn variant in TTR.The binocular visual acuity recovered to 1.0 after binocular vitrectomy.CONCLUSION Vitreous amyloidosis is rare in the clinic and gene testing can assist the diagnosis accurately and effectively.

Rare manifestation of familial vitreous amyloidosis caused by Gly103Arg transthyretin

AIM:To identify and analyze the genotype of the patients with special ocular manifestations of familial vitreous amyloidosis(FVA)in a Chinese Han family.METHODS:Pars plana vitrectomy(PPV)surgery was performed on a 52-year-old Chinese woman presented with vitreous amyloidosis and progressive visual impairment,without evidence of cardiac,renal,gastrointestinal,central nervous system or peripheral nervous system dysfunction.During the surgery,the patient presented with a gray-white dense and thick cotton woollike change in the vitreous body,accompanied by complete retinal detachment.Additionally,hard,free and movable yellow-white deposits were observed in the posterior pole and surrounding retina,the vitreous and subretinal deposits were examined by Congo red staining and immunohistochemical pathological examination,and whole exome sequencing was performed on blood samples from the patient and her cousin.RESULTS:During the operation,it was discovered that there was a complete detachment of the retina and a significant amount of hard,free-floating yellow-white deposits were observed beneath the posterior pole and surrounding retina.This is an exceedingly rare ocular manifestation.Pathological examination of the vitreous and subretinal deposit specimens revealed positive Congo red staining,as well as elevated vascular endothelial growth factor(VEGF)expression in vascular endothelial cells within the sediment specimens upon immunohistochemical examination.The patient and her cousin both exhibited a heterozygous mutation in Glyl03Arg within the transthyretin(TTR)gene,resulting in a substitution of glycine(Gly)at position 103 with arginine(Arg).CONCLUSION:FVA may present with various ocular manifestations,but panretinal detachment is a rare occurrence.In cases where retinal detachment persists for an extended period of time,amyloid deposits may form under the retina through retinal tears,leading to subretinal deposits that can impede retinal reattachment and negatively impact visual prognosis.Elevated levels of VEGF in the eyes of FVA patients may indicate an overexpression state,necessitating careful postoperative follow-up.The heterozygous mutation Gly103Arg may represent a unique pathogenic site in Chinese individuals.

Elevated interleukin-6 levels are associated with impaired outcome in cardiac transthyretin amyloidosis

BACKGROUND Elevated interleukin(IL)-6-levels have been described in familial variant transthyretin amyloidosis(ATTRv)associated polyneuropathy and heart failure.However,IL-6 in cardiac ATTR amyloidosis(ATTR-CM)and its prognostic value have not been investigated yet.AIM We aim to study the correlation between IL-6 levels with clinical presentation(Gillmore-class)and outcome[heart transplantation or death(htx/death)],or the combined endpoint of cardiac decompensation or htx/death in ATTR-CM.METHODS IL-6 levels of 106 ATTR-CM patients[54 wild-type ATTRwt,52 ATTRv-CM],15 asymptomatic carriers of ATTR mutations(aATTRv-CM)and 27 healthy donors were quantified using Luminex technology.Statistical analysis was performed using parametric survival regression models.RESULTS We found that IL-6 levels from wild-type ATTR patients were significantly elevated compared to healthy controls,while aATTRv-CM carriers and ATTRv-CM patients did not show a significant difference.IL-6 levels showed significantly higher values in increasing Gillmore classes.Univariate analyses revealed association of low IL-6 levels with cardiac decompensation and htx/death[odds ratio:0.26(0.09-0.72),P=0.01]and htx/death[odds ratio:0.15(0.04-0.58),P=0.006].However,in the multivariate model,no significant improvement of risk prediction was seen for IL-6,while established prognostic factors were significantly associated with outcome.CONCLUSION Raised IL-6 levels correlate with clinical presentation and are associated with worse outcome in ATTR-CM but do not improve stratification in addition to established risk factors.

Screening for Transthyretin Cardiac Amyloidosis in Patients with Bilateral Carpal Tunnel Syndrome: Identifying Missed Opportunities for Early Detection and Treatment

Purpose: Transthyretin cardiac amyloidosis (ATTR-CA) has been linked to many extra-cardiac manifestations including bilateral carpal tunnel syndrome (CTS). The aim of this study is to analyze patients with bilateral CTS to identify patients with high-risk features or “red flags” for ATTR-CA, identify if systematic screening was done for ATTR-CA and define opportunities for improved detection. Methods: Out of >5000 patients with bilateral CTS evaluated in a single tertiary care center in Southeast Michigan (2010-2016), we retrospectively studied a focused population of patients: men > 50 years and women > 60 years old with bilateral CTS and atrial fibrillation (n = 295). Baseline demographic, comorbidities, and electrocardiographic and echocardiographic findings were analyzed. A high-risk group suspicious for ATTR-CA was identified as patients with bilateral CTS, atrial fibrillation, and concomitant “red flags” including heart failure and left ventricular hypertrophy. Results: Out of 295 patients, 51.2% were female, 75.6% were White, and 22.4% were African American. Upon comparing the high-risk group (n = 67) with the remaining study population (n = 228), both diagnosis of ATTR-CA and mortality were higher among the high-risk group (7.5% vs 0.4% and 43.3% vs 24.6%, respectively, P = 0.003). Conclusions: A substantial number of bilateral CTS patients had additional “red flags” warranting formal evaluation for ATTR-CA;however, systematic evaluation for cardiac amyloidosis was not performed in many patients. This emphasizes that Multidisciplinary collaboration is needed to create a systematic workflow and to raise awareness amongst cardiologists and other physicians for suspecting ATTR-CA in bilateral CTS patients who have additional “red flags”.

Hereditary Transthyretin Amyloidosis in Eight Chinese Families

Background： Mutations of transthyretin （TTR） cause the most common type of autosomal-dominant hereditary systemic amyloidosis, which occurs worldwide. To date, more and more mutations in the TTR gene have been reported. Some variations in the clinical presentation are often observed in patients with the same mutation or the patients in the same family. The purpose of this study was to find out the clinicopathologic and genetic features of Chinese patients with hereditary TTR amyloidosis. Methods： Clinical and necessary examination materials were collected from nine patients of eight families with hereditary TTR amyloidosis at Peking University First Hospital from January 2007 to November 2014. Sural nerve biopsies were taken for eight patients and skin biopsies were taken in the calf/upper arm for two patients, for light and electron microscopy examination. The TTR genes from the nine patients were analyzed. Results： The onset age varied from 23 to 68 years. The main manifestations were paresthesia, proximal and/or distal weakness, autonomic dysfunction, cardiomyopathy, vitreous opacity, hearing loss, and glossohypertrophia. Nerve biopsy demonstrated severe loss ofmyelinated fibers in seven cases and amyloid deposits in three. One patient had skin amyloid deposits which were revealed from electron microscopic examination. Genetic analysis showed six kinds of mutations of TTR gene, including Val30Met, Phe33Leu, Ala36Pro, Val30Ala, Phe33Val, and Glu42Gly in exon 2. Conclusions： Since the pathological examinations ofsural nerve were negative for amyloid deposition in most patients, the screening for TTR mutations should be performed in all the adult patients, who are clinically suspected with hereditary TTR amyloidosis.

Clinical and laboratory characteristics of patients having amyloidogenic transthyretin deposition in osteoarthritic knee joints

Objective： Our aim was to investigate clinical and laboratory characteristics of osteoarthritic patients who had amyloid deposition in their knee joints. Methods： Synovial membranes were obtained from 36 patients with knee osteoarthritis （OA） who underwent joint replacement surgery. From this sample, the diagnosis of amyloid was deter- mined by Congo red staining, which demonstrated apple-green birefringence under a polarized microscope. All syn- ovial membranes were immunohistochemically characterized for the expressions of amyloid immunoglobulin light chain （AL-K and AL-,k）, serum amyloid-A （SAA）, amyloidogenic transthyretin （ATTR）, and amyloidogenic 152- microglobulin （A152M）. Matrix-assisted laser desorption-ionizaton/time of flight mass spectrometry （MALDI-TOF MS） was used to analyze transthyretin （TTR） isoforms in the serum of each patient. Results： Nine cases （25%） were found to be amyloid-positive. Immunohistochemicaliy, eight cases （88.9%） had ATTR deposition, and one sample （11.1%） was shown to be AL-K-positive. MALDI-TOF MS identified that the TTR in the serum of the patients was unmodified wild-type TTR, TTR-Cys-S-S-Cys, and TTR-Cys-S-S-CysGly. The age at surgery and the disease duration were sig- nificantly higher in the ATTR-positive group than in the ATTR-negative group. Knee score and function score were significantly lower in the ATTR-positive group than in the ATTR-negative group. Conclusions： Amyloid deposition in synovial membranes of OA patients was found to be ATTR and AL-K. TTR in the serum of the patients was unmodified wild-type TTR together with two isoforms. The high age at surgery, long disease duration, and a deteriorated knee function were associated with ATTR amyloid deposition in the osteoarthritic knee joints.

Recent advances in transthyretin amyloidosis therapy

Mutant(MT)forms of transthyretin(TTR)cause the most common type of autosomal-dominant hereditary systemic amyloidosis—familial amyloidotic polyneuropathy(FAP).Until 20 years ago,FAP was thought to be an endemic disease,but FAP is known to occur worldwide.To date,more than 130 mutations in the TTR gene have been reported.Genotype-phenotype correlations are seen in FAP,and some variation in clinical presentation is often observed in individual kindreds with the same mutation and even among family members.Of the pathogenic TTR mutations,Val30Met was the first to be identified and is the most frequent known mutation found throughout the world.Studies of patients with FAP amyloidogenic TTR(ATTR)Val30Met documented sensorimotor polyneuropathy,autonomic dysfunction,heart and kidney failure,gastrointestinal tract(GI)disorders,and other symptoms leading to death,usually within 10 years of the onset of disease.Diagnosis is sometimes delayed,especially in patients without a clear family history and typical clinical manifestations,since diagnosis requires various studies and techniques such as histopathology,genetic testing,and mass spectrometry.For treatment of FAP,liver transplantation(LT)reportedly halts the progression of clinical manifestations.Exchange of an FAP patient’s diseased liver with a healthy liver causes MT TTR in the body to be replaced by wild-type(WT)TTR.Although clinical evaluations indicated that progression of other clinical symptoms such as peripheral neuropathy,GI symptoms,and renal involvement usually halted after LT in FAP ATTR Val30Met patients,recent studies suggested that LT failed to prevent progression of cardiac amyloidosis in FAP ATTR Val30Met patients after LT,with this failure reportedly being due to continued formation of amyloid that derived mainly from WT TTR secreted from the transplanted non-mutant liver graft.In recent years,many therapeutic strategies have been proposed,and several ongoing therapeutic trials involve,for example,stabilizers of TTR tetramers(tafamidis and diflunisal)and gene therapies to suppress TTR expression(antisense methods and use of small interfering RNAs).These novel therapies may prove to prevent progression of FAP.

骨性关节病关节组织发现Transthyretin和ImmunoglobulinLightChain两种类型淀粉样变

目的 分析老年性关节病组织内是否存在淀粉样变沉积,为解析导致淀粉样变形成机制奠定基础。方法 选择骨关节病进行手术治疗患者88例,男46例,50-74岁,女42例,53-72岁;同时作为对照组筛选外伤关节手术患者14例,男8例,年龄58-75岁,女6例,年龄57-77岁。留取关节组织进行刚果红染色,结合偏振光观察组织内是否存在淀粉样变。另外,利用组织标本进行immunoglobulin light chain（AL-kappa）、immunoglobulin light chain（AL-lambda）、transthyretin（TTR）、2-microglobulin（2-M）和serum amyloid A（SAA）五种抗体免疫组织化学染色。利用Matrix-assisted laser desorption-ionizaton/time of flight mass spectrometry（MALDI-TOF MS）系统分析TTR基因突变型。结果 在骨关节病患者关节组织内发现Congo Red染色阳性并且可见偏振光显微镜绿色荧光,阳性率为26.1%（23/88）,利用TTR、SAA、免疫球蛋白轻链等五种抗体进行免疫组织化学染色,发现有TTR和AL-kappa两种淀粉样变蛋白,其发生率为TTR22.7%,AL-kappa为3.4%,且TTR基于型均未发现有突变型。结论发现老年性骨关节病组织内出现淀粉样变,免疫组织化学方法发现TTR和AL-kappa两种类型相关淀粉样变。

Molecular Dynamics Simulations and Steered Molecular Dynamics Simulations of Glabridin Bound to Wild Type and V30A Mutant Transthyretin： Ligand-linked Perturbation of Tertiary Conformation

Transthyretin（TTR）, as a tetrameric protein, functions as a neuroprotector. The native TTR homotetramer dissociates into dimers and monomers. Dimers and monomers self-assemble into amyloid fibrils, and this process can lead to some diseases. Native TTR homotetramer is a widely accepted model for TTR amyloid formation. In this study, simulations using molecular dynamics（MD） and steered MD（SMD） were performed to explore the mechanisms for glabridin（Glab）, a specific inhibitor for TTR binding, for V30A mutant and wild-type（WT） TTR. MD simulation results indicate that, compared with Glab binding to WT and V30A mutant, the WT TTR could lead to the collapse of β-strands from Ser52 to His56 at chain A. This phenomenon facilitated the easy dissociation of chains A and C. Calculations of the binding free energy between the two chains showed that the V30A-Glab TTR complex displayed a lower binding energy than other systems（WT TTR and WT-Glab TTR）. Then, SMD simulation was performed to ex- plore the unbinding pathway for Glab through the WT and V30A mutant TTR. The results show that Lysl 5（chain A） produced a hydrogen bond with Glab at the force peak via the WT TTR tunnel. Meanwhile, in the V30A TTR mutant, the hydrogen bond between Lysl 5（chain A） and Glab was broken at the force peak. This condition was beneficial for Glab to be taken off from the protein. Our theoretical results will be useful in designing a new specific inhibitor of TTR protein to control the TTR homotetramer dissociation.

3D-QSAR and docking studies on 2-arylbenzoxazole and linker-Y transthyretin amyloidogenesis inhibitors

Transthyretin(TTR),a plasma protein with a tetramer structure,could form amyloid fibril associated with several human diseases through the dissociation of tetramer and the misfolding of monomer.These amyloidogenesis can be inhibited by small molecules which bind to the central channel of TTR.A number of small molecules like 2-arylbenzoxazoles(ABZ)analogues are proposed as promising therapeutic strategy to treat amyloidosis.In this work,comparative molecular field analysis(CoMFA)and comparative molecular similarity indices analysis(CoMSIA)three-dimensional quantitative structure-activity relationship(3D-QSAR)and docking studies were performed on series of 2-arylbenzoxazoles(ABZ)and linker-Y analogues to investigate the inhibitory activities of TTR amyloidogenesis at atomic level.Significant correlation coefficients for ABZ series(CoMFA,r2=0.877,q2=0.431;CoMSIA,r2=0.836,q2=0.447)and those for linker-Y series(CoMFA,r2=0.828,q2=0.522;CoMSIA,r2=0.800,q2=0.493)were obtained,and the generated models were validated using test sets.In addition,docking studies on 6 compounds binding to TTR were performed to analyze the forward or reverse binding mode and interactions between molecules and TTR.These results from 3D-QSAR and docking studies have great significance for designing novel TTR amyloidogenesis inhibitors in the future.