32例噬血细胞综合征实验室检查及临床分析

目的分析继发性噬血细胞综合征(HPS)患者的实验室指标及临床特点,以提高对该病的认识。方法对2005年6月~2015年6月在广西肿瘤防治研究所治疗的32例继发性HPS患者的病因、临床表现、实验室特征、治疗及预后等进行分析。结果 32例患者中,血液系统肿瘤17例,单纯感染13例,自身免疫性疾病2例。临床上以发热(100%)、肝脾(87.5%)及淋巴结(43.8%)肿大为主要表现。实验室检查以血细胞减少(100%)、铁蛋白升高(96.9%)、自然杀伤细胞活性降低或缺失(96.9%)为主,纤维蛋白原降低(68.8%),甘油三酯升高(56.3%),81.3%的患者骨髓涂片检查可见噬血细胞现象。结论继发性HPS可由多种病因引起,临床表现多样,对可疑患者及时进行相关实验室检查有助于早期明确诊断;合并多脏器衰竭、DIC的患者预后不良。     

Genetic variants and mutational spectrum of Chinese Hermansky–Pudlak syndrome patients

Hermansky–Pudlak syndrome (HPS) is a rare recessive disorder characterized by oculocutaneous albinism or ocular albinism, bleeding diathesis, and other symptoms such as colitis and pulmonary fibrosis. Eleven causative genes have been identified for HPS‐1–HPS‐11 subtypes in humans. We have identified 16 newly reported patients including the first HPS‐2 case in the Chinese population. In a total of 40 HPS patients, hypopigmentation was milder in HPS‐3, HPS‐5, and HPS‐6 patients than in HPS‐1 and HPS‐4 patients. HPS‐1 accounted for 47.5% (19 of 40) of HPS cases which is the most common subtype. Exons 11 and 19 were the hotspots of the HPS1 gene mutations. In total, 55 allelic variants were identified in HPS1–HPS6 gene, of which 17 variants were previously unreported. These results will be useful for the evaluation of the relationship between HPS genotypes and phenotypes, and for the precise intervention of HPS patients in the Chinese population.     

Clinico‐molecular analysis of eleven patients with Hermansky–Pudlak type 5 syndrome,a mild form of HPS

Hermansky–Pudlak syndrome (HPS), first described in 1959, is a rare form of syndromic oculocutaneous albinism associated with bleeding diathesis and in some cases pulmonary fibrosis and granulomatous colitis. All 10 HPS types are caused by defects in vesicle trafficking of lysosome‐related organelles (LRO) proteins. The HPS5 protein associates with HPS3 and HPS6 to form the biogenesis of lysosome‐related organelles complex‐2 (BLOC‐2). Here, we report the clinical and genetic data of 11 patients with HPS‐5 analyzed in our laboratory. We report 11 new pathogenic variants. The 11 patients present with ocular features that are typical for albinism, with mild hypopigmentation, and with no other major complication, apart from a tendency to bleed. HPS‐5 therefore appears as a mild form of HPS, which is often clinically undistinguishable from mild oculocutaneous or ocular forms of albinism. Molecular analysis is therefore required to establish the diagnosis of this mild HPS form, which has consequences in terms of prognosis and of clinical management of the patients.     

Characterization of melanosomes and melanin in Japanese patients with Hermansky–Pudlak syndrome types 1, 4, 6, and 9

Hermansky–Pudlak syndrome (HPS) is an autosomal recessive disorder characterized by oculocutaneous albinism (OCA), a bleeding tendency, and ceroid deposition. Most of the causative genes for HPS encode subunits of the biogenesis of lysosome‐related organelles complex (BLOC). In this study, we identified one patient each with HPS4, HPS6, and HPS9 by whole‐exome sequencing. Next, we analyzed hair samples from the three patients and representative patients with HPS1 and controls using electron microscopy and chemical methods. All HPS patients had fewer, smaller, and more immature melanosomes than healthy controls. Further, all patients showed reduced total melanin content and increased levels of benzothiazine‐type pheomelanin. The results of this study demonstrate the impact of the dysfunctions of BLOCs on the maturation of melanosomes and melanin levels and composition through analysis of their hair samples.     

Enhanced production of lactic acid through the use of a novel aqueous two-phase system as an extractive fermentation system

 In order to enhance the productivity of lactic acid and reduce the end-product inhibition of fermentation, the partitioning and growth of four different strains of lactic acid bacteria in three different aqueous two-phase systems were studied. Polyethyleneglycol/ dextran, polyethyleneglycol/hydroxypropyl starch polymer (HPS), and a random copolymer of ethylene oxide and propylene oxide (EO-PO)/HPS were used as polymer systems. One strain each of Lactococcus lactis subsp. lactis and of Lactobacillus delbrueckii subsp. delbrueckii partitioned completely to the interface and bottom phase in two-phase systems with low polymer concentrations of EO-PO/HPS100 and EO-PO/ HPS200. The growth and production of lactic acid by two of three L. lactis strains in a two-phase system with 5.5% (w/w) EO-PO and 12.0% (w/w) HPS100 were reduced by less than 10% compared with a reference fermentation in a normal growth medium. The viability of L. lactis subsp. lactis ATCC 19435 was maintained for at least 50 h and with four top-phase replacements during extractive fermentation in the EO-PO/HPS100 system. Moreover, when cell density reached the stationary phase in the first extractive fermentation, the lactate production in this aqueous two-phase system was maintained. Received: 2 October 1995/Received revision: 16 January 1996/Accepted: 22 January 1996     

安徽PRSS发病猪副猪嗜血杆菌的分离鉴定及其耐药性分析

目的了解猪繁殖与呼吸综合征（PRRS）发病猪继发感染副猪嗜血杆菌（HPS）的情况及分离菌株的药物感受性。方法应用细菌分离培养、形态学检查、生化试验和PCR技术,对2007年1月至2008年12月从安徽不同地区采集的146头份PRRS发病猪的病料进行HPS检测,并采用标准K-B纸片法对分离菌株进行14种抗菌药物敏感试验。结果分离鉴定出12株HPS,检出率为8.22%（12/146）;12株HPS对氯霉素100%敏感,环丙沙星为91.7%,阿莫西林和新霉素为83.3%,对罗红霉素100%耐药,阿米卡星为83.3%。结论安徽省不同地区PRRS感染猪群中均存在程度不同的HPS感染,各地区HPS分离株表现出形态上的变化特征和一致的生化特性,且有对临床常用抗菌药物耐药性增强的趋势。     

NGS‐based 100‐gene panel of hypopigmentation identifies mutations in Chinese Hermansky–Pudlak syndrome patients

Hermansky–Pudlak syndrome (HPS) is a rare recessive disorder characterized by hypopigmentation, bleeding diathesis, and other symptoms due to multiple defects in lysosome‐related organelles. Ten HPS subtypes have been identified with mutations in HPS1 to HPS10. Only four patients with HPS‐1 have been reported in Chinese population. Using next‐generation sequencing (NGS), we have screened 100 hypopigmentation genes and identified four HPS‐1, two HPS‐3, one HPS‐5, and three HPS‐6 in Chinese HPS patients with typical ocular or oculocutaneous albinism and the absence of platelet dense granules together with other variable phenotypes. All these patients except one homozygote were compound heterozygotes. Among these mutations, 14 were previously unreported alleles (four in HPS1, three in HPS3, two in HPS5, five in HPS6). Our results demonstrate the feasibility and utility of NGS‐based panel diagnostics for HPS. Genotyping of HPS subtypes is a prerequisite for intervention of subtype‐specific symptoms.     

Instability of BLOC‐2 and BLOC‐3 in Chinese patients with Hermansky‐Pudlak syndrome

Hermansky‐Pudlak syndrome (HPS) is a rare recessive disorder characterized by oculocutaneous albinism (OCA) or ocular albinism (OA), bleeding tendency, and other symptoms due to multiple defects in tissue‐specific lysosome‐related organelles. Ten HPS subtypes have been characterized with mutations in HPS1 to HPS10, which encode the subunits of BLOC‐1, ‐2, ‐3, and AP‐3. Using next‐generation sequencing (NGS), we have screened 100 hypopigmentation genes in OCA or OA patients and identified four HPS‐1, one HPS‐3, one HPS‐4, one HPS‐5, and three HPS‐6. The HPS‐4 case is the first report in the Chinese population. Among these 20 mutational alleles, 16 were previously unreported alleles (6 in HPS1, 1 in HPS3, 2 in HPS4, 2 in HPS5, and 5 in HPS6). BLOC‐2 and BLOC‐3 were destabilized due to the mutation of these HPS genes which are so far the only reported causative genes in Chinese HPS patients, in which HPS‐1 and HPS‐6 are the most common subtypes. The mutational spectrum of Chinese HPS is population specific.     

A clinical variant of familial Hermansky-Pudlak syndrome

Hermansky-Pudlak syndrome (HPS) is an autosomal recessive inherited disease consisting of (1) partial oculocutaneous albinism (with nystagmus, strabism, and visual acuity loss), (2) platelet storage pool deficiency (with bleeding diathesis), and (3) disorder of "ceroid" metabolism with a multisystem tissue lysosomal ceroid deposition. HPS is less uncommon in Puerto Rico, where the most important studies have been performed, but is a very rare disease in Europe. HPS basic defect remains unknown, even if an HPS-causing gene was identified in chromosome segment 10q23-q23.3, and several mutations have been reported. The aim of this article is to discuss, on the basis of a review of relevant literature, a new familial HPS clinical variant observed in 2 young sisters (aged 16 and 23 years old, respectively), characterized by the typical symptoms of this syndrome. Our patients also suffered from diffuse interstitial pulmonary disease and an unexpectedly increased platelet aggregation and were prone to bacterial infections. Interestingly, we observed urinary tract abnormality in the younger HPS sister and a porencephalic cyst in the older HPS sister; both of these developmental defects have been reported in the Cross syndrome (or oculocerebral hypopigmentation syndrome). It seems that in our patients, an overlapping of the phenotypic manifestations of different rare syndromes may be present. The presence of ceroid-like autofluorescent material in urinary sediment together with the histologic aspects and the autofluorescence of oral mucosa biopsy are consistent with a ceroid-like lipofuscin storage. HPS should be carefully tested for in suspected cases to prevent the severe visual impairment, rapidly progressive pulmonary fibrosis, and other complications associated with this disorder.     

37例儿童噬血细胞综合征的临床分析

目的:分析儿童噬血细胞综合征(hemophagocytic syndrome,HPS)的病因、临床表现、实验室检查结果、治疗和预后特点。方法:回顾性分析我院收治的37例HPS患儿的临床资料。结果:37例HPS患儿(男24例、女13例),年龄2月~9岁,5例(13.5%)有明显家族史,获得性HPS32例(86.5%),包括EB病毒感染16例、巨细胞病毒感染7例,其他原因9例;所有患儿均表现为发热,肝脾肿大,外周血白细胞、血红蛋白、血小板、白蛋白、纤维蛋白原减低,TG、ALT、AST、LDH、铁蛋白升高;5例遗传性HPS患儿死亡4例,放弃治疗1例,剩余32例患儿中好转20例(62.5%),包括痊愈17例,完全缓解后继续治疗中3例,未好转12例(37.5%),其中死亡7例,病情危重放弃治疗3例,复发2例。12例未好转病例中,9例为EBV感染,1例为肾母细胞瘤,1例为幼年类风湿性关节炎合并CMV感染,1例原因不明。遗传性HPS的好转率较继发性HPS明显降低,差异有统计学意义(X2=5.30,P0.05),继发性HPS中EBV感染者的好转率较非EBV感染者低,差异有统计学意义(X2=4.80,P0.05)。结论:及时诊断儿童HPS并明确其病因,对该病的治疗及预后具有重要意义。     

Hantaviruses: molecular biology, evolution and pathogenesis

Hantaviruses are tri-segmented negative sense single stranded RNA viruses that belong to the family Bunyaviridae. In nature, hantaviruses are exclusively maintained in the populations of their specific rodent hosts. In their natural host species, hantaviruses usually develop a persistent infection with prolonged virus shedding in excreta. Humans become infected by inhaling virus contaminated aerosol. Unlike asymptomatic infection in rodents, hantaviruses cause two acute febrile diseases in humans: hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). The mortality rate varies from 0.1% to 40% depending on the virus involved. Hantaviruses are distributed world wide, with over 150,000 HFRS and HPS cases being registered annually. In this review we summarize current knowledge on hantavirus molecular biology, epidemiology, genetic diversity and co-evolution with rodent hosts. In addition, special attention was given in this review to describing clinical manifestation of HFRS and HPS, and advances in our current understanding of the host immune response, treatment, and prevention.     

Human Albinism and Animal Models of Albinism

Ten phenotypic forms of oculocutaneous albinism (OCA) and four forms of ocular albinism (OA) have been identified in man. All have optic neuronal decussation defects at the optic chiasm. Thus any proposed animal model for these disorders must share optic neuronal decussation defects in addition to hypopigmentation. Three, tyrosinase-negative (ty-neg), yellow mutant (ym), and platinum (pt), OCA appear to be allelic in humans. Two, ty-neg and pt, OCA appear to be analogous to c-locus mutants c/c and c^p/c^p in mice, but no homologue is known in mice for ym OCA. Tyrosinase-positive (ty-pos) OCA, which is nonallelic with ty-neg OCA, shares many morphological and biochemical features with pink-eyed mice. Chediak-Higashi syndrome (CHS) and Hermansky-Pudlak syndrome (HPS) appear to be due to genes acting extrinsic to the melanin pathway. CHS is homologous with beige in mice. HPS was investigated in northwestern Puerto Rico, where it affects approximately 1 in 2,000 persons. Approximately 68% of 37 deceased HPS patients died from sequelae of ceroid storage disease, restrictive lung disease between ages 35 and 46 years (43%), and granulomatous colitis (8%) or hemorrhage (16%). The most accurate and consistent diagnostic feature of HPS is lack of platelet dense bodies. HPS patients with ceroid storage disease had high urinary levels of long-chain isoprenoid alcohols, dolichols, similar to that seen in the neuronal-ceroid lipofuscinoses (Batten disease). Dolichols are constituents of lysosomes, and their elevation in HPS suggests that this syndrome carries a lysosomal defect. There is no degradative pathway for ceroid and dolichols, which are eliminated by exocytosis. The exocytic process is thought to involve a thioendoproteinase. Pale-ear mice have been proposed as a model for HPS; their platelets lack dense bodies, and they are depigmented. Leupeptin, a thioendoproteinase inhibitor, administered to 100-day-old pale-eared and black wild-type C57 mice for 10 days resulted in the accumulation of ceroid in tissues in the same pattern as that in HPS, but granulomas of gut or fibrosis of lungs were not seen. Determinations of homology between mice and men at the molecular level is now possible with the isolation of mouse tyrosinase by Yamamoto et al. and isolation by Kwon et al. of human tyrosinase mapping at the c-locus in mice.     

Second external quality assurance study for the serological diagnosis of hantaviruses in Europe

Hantaviruses are endemic throughout the world and hosted by rodents and insectivores. Two human zoonoses, hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS), are caused by hantaviruses and case fatality rates have reached 12% for HFRS and 50% for HPS in some outbreaks. Symptomatic hantavirus infections in Europe are summarised as HFRS mainly due to Puumala, Dobrava-Belgrade and Saaremaa virus. While HFRS has an overall low incidence in Europe, the number of cases varies from 100 per year in all Eastern and Southern Europe up to 1,000 per year only in Finland. To assess the quality of hantavirus diagnostics, the European Network for the Diagnostics of "Imported" Viral Diseases (ENIVD) organised a first external quality assurance (EQA) in 2002. The purpose of this second EQA study is to collect updated information on the efficiency and accurateness of hantavirus serological methods applied by expert laboratories. A serum panel of 14 samples was sent to 28 participants in Europe of which 27 sent results. Performance in hantavirus diagnosis varied not only on the method used but also on the laboratories and the subclass of antibodies tested. Commercial and in-house assays performed almost equally. Enzyme immunoassays were mainly used but did not show the best performances while immunoblot assays were the less employed and showed overall better performances. IgM antibodies were not detected in 61% of the positive IgM samples and IgM detection was not performed by 7% of the laboratories indicating a risk of overlooking acute infections in patients. Uneven performances using the same method is indicating that there is still a need for improving testing conditions and standardizing protocols.     

BLOC-2 subunit HPS6 deficiency affects the tubulation and secretion of von Willebrand factor from mouse endothelial cells

Hermansky-Pudlak syndrome(HPS) is a recessive disorder with bleeding diathesis, which has been linked to platelet granule defects. Both platelet granules and endothelial Weibel-Palade bodies(WPBs)are members of lysosome-related organelles(LROs) whose formation is regulated by HPS protein associated complexes such as BLOC(biogenesis of lysosome-related organelles complex)-1,-2,-3, AP-3(adaptor protein complex-3) and HOPS(homotypic fusion and protein sorting complex). Von Willebrand factor(VWF) is critical to hemostasis, which is stored in a highly-multimerized form as tubules in the WPBs. In this study, we found the defective, but varying, release of VWF into plasma after desmopressin(DDAVP) stimulation in HPS1(BLOC-3 subunit), HPS6(BLOC-2 subunit), and HPS9(BLOC-1 subunit)deficient mice. In particular, VWF tubulation, a critical step in VWF maturation, was impaired in HPS6 deficient WPBs. This likely reflects a defective endothelium, contributing to the bleeding tendency in HPS mice or patients. The differentially defective regulated release of VWF in these HPS mouse models suggests the need for precise HPS genotyping before DDAVP administration to HPS patients.     

Molecular characterization of the protein encoded by the Hermansky-Pudlak syndrome type 1 gene

Hermansky-Pudlak syndrome (HPS) comprises a group of genetic disorders characterized by defective lysosome-related organelles. The most common form of HPS (HPS type 1) is caused by mutations in a gene encoding a protein with no homology to any other known protein. Here we report the identification and biochemical characterization of this gene product, termed HPS1p. Endogenous HPS1p was detected in a wide variety of human cell lines and exhibited an electrophoretic mobility corresponding to a protein of approximately 80 kDa. In contrast to previous theoretical analysis predicting that HPS1p is an integral membrane protein, we found that this protein was predominantly cytosolic, with a small amount being peripherally associated with membranes. The sedimentation coefficient of the soluble form of HPS1p was approximately 6 S as inferred from ultracentrifugation on sucrose gradients. HPS1p-deficient cells derived from patients with HPS type 1 displayed normal distribution and trafficking of the lysosomal membrane proteins, CD63 and Lamp-1. This was in contrast to cells from HPS type 2 patients, having mutations in the beta3A subunit of the AP-3 adaptor complex, which exhibited increased routing of these lysosomal proteins through the plasma membrane. Similar analyses performed on fibroblasts from 10 different mouse models of HPS revealed that only the AP-3 mutants pearl and mocha display increased trafficking of Lamp-1 through the plasma membrane. Taken together, these observations suggest that the product of the HPS1 gene is a cytosolic protein capable of associating with membranes and involved in the biogenesis and/or function of lysosome-related organelles by a mechanism distinct from that dependent on the AP-3 complex.     

Role of mixed Th1 and Th2 serum cytokines on pathogenesis and prognosis of hantavirus pulmonary syndrome

The hantavirus pulmonary syndrome (HPS) is an emerging syndrome in the Americas. The disease results from intense immune activation and changes in vascular permeability. The aim of this study was to determine the profile of serum cytokines in HPS patients looking for correlation with the clinical parameters, severity and outcome of illness. Studying 21 HPS patients, we found that IL-6 may have an important role in the pathogenesis of HPS, being associated with fatal outcome. Our results also support a mixed Th1/Th2 immune response during the course of HPS and that the magnitude of Th1 response effector cytokines is correlated to HPS severity. The decreased levels of TGF-beta observed in HPS patients suggest that immunoregulatory activity could be damaged in these patients.     

Aberrant lung structure,composition, and function in a murine model of Hermansky-Pudlak syndrome

Hermansky-Pudlak syndrome (HPS) is a genetically heterogeneous inherited disease causing hypopigmentation and prolonged bleeding times. An additional serious clinical problem of HPS is the development of lung pathology, which may lead to severe lung disease and premature death. No cure for the disease exists, and previously, no animal model for the HPS lung abnormalities has been reported. A mouse model of HPS, which is homozygously recessive for both the Hps1 (pale ear) and Hps2 (pearl) genes, exhibits striking abnormalities of lung type II cells. Type II cells and lamellar bodies of this mutant are greatly enlarged, and the lamellar bodies are engorged with surfactant. Mutant lungs accumulate excessive autofluorescent pigment. The air spaces of mutant lungs contain age-related elevations of inflammatory cells and foamy macrophages. In vivo measurement of lung hysteresivity demonstrated aberrant lung function in mutant mice. All these features are similar to the lung pathology described in HPS patients. Morphometry of mutant lungs indicates a significant emphysema. These mutant mice provide a model to further investigate the lung pathology and therapy of HPS. We hypothesize that abnormal type II cell lamellar body structure/function may predict future lung pathology in HPS.     

Lessons of a day hospital: Comprehensive assessment of patients with albinism in a European setting

Albinism is a rare genetic disease, comprising syndromic and non‐syndromic forms. We assessed clinical and genetic characteristics in a prospective evaluation of 64 patients (33 children and 31 adults) seen at a specialized day hospital. Causative genetic mutations were found in TYR (23/64, 35.9%), OCA2 (19/64, 29.7%), TYRP1 (1/64, 1.6%), SLC45A2 (12/64, 18.7%), C10orf11 (1/64, 1.6%), HPS1 (3/64, 4.7%), HPS5 (1/64, 1.5%), HPS6 (1/64, 1.6%) and GPR143 (2/64, 3.1%). Causative mutations remained undetermined for one patient (1.6%). Heterogeneity for hair and skin phenotype was noted across and within the different genotypes. Skin and hair hypopigmentation did not correlate with visual impairment. The diagnosis of unrecognized syndromic forms and of cases of ocular albinism in this prospective and comprehensive series of patients with albinism in a European setting is remarkable. Photoprotection was overall good but not optimal.     

Characterization of BLOC-2, a complex containing the Hermansky-Pudlak syndrome proteins HPS3, HPS5 and HPS6

Hermansky–Pudlak syndrome (HPS) defines a group of at least seven autosomal recessive disorders characterized by albinism and prolonged bleeding due to defects in the lysosome-related organelles, melanosomes and platelet-dense granules, respectively. Most HPS genes, including HPS3, HPS5 and HPS6 , encode ubiquitously expressed novel proteins of unknown function. Here, we report the biochemical characterization of a stable protein complex named Biogenesis of Lysosome-related Organelles Complex-2 (BLOC-2), which contains the HPS3, HPS5 and HPS6 proteins as subunits. The endogenous HPS3, HPS5 and HPS6 proteins from human HeLa cells coimmunoprecipitated with each other from crude extracts as well as from fractions resulting from size-exclusion chromatography and density gradient centrifugation. The native molecular mass of BLOC-2 was estimated to be 340 ± 64 kDa. As inferred from the biochemical properties of the HPS6 subunit, BLOC-2 exists in a soluble pool and associates to membranes as a peripheral membrane protein. Fibroblasts deficient in the BLOC-2 subunits HPS3 or HPS6 displayed normal basal secretion of the lysosomal enzyme β-hexosaminidase. Our results suggest a common biological basis underlying the pathogenesis of HPS-3, -5 and -6 disease.