CHCHD2 Thr61Ile mutation impairs F1F0-ATPase assembly in in vitro and in vivo models of Parkinson's disease

Mitochondrial dysfunction is a significant pathological alte ration that occurs in Parkinson's disease(PD),and the Thr61lle(T61I)mutation in coiled-coil helix coiled-coil helix domain containing 2(CHCHD2),a crucial mitochondrial protein,has been reported to cause Parkinson's disease.FIFO-ATPase participates in the synthesis of cellular adenosine triphosphate(ATP)and plays a central role in mitochondrial energy metabolism.However,the specific roles of wild-type(WT)CHCHD2 and T611-mutant CHCHD2 in regulating F1FO-ATPase activity in Parkinson's disease,as well as whether CHCHD2 or CHCHD2 T61I affects mitochondrial function through regulating F1FO-ATPase activity,remain unclea r.Therefore,in this study,we expressed WT CHCHD2 and T61l-mutant CHCHD2 in an MPP^(+)-induced SH-SY5Y cell model of PD.We found that CHCHD2 protected mitochondria from developing MPP^(+)-induced dysfunction.Under normal conditions,ove rexpression of WT CHCHD2 promoted F1FO-ATPase assembly,while T61I-mutant CHCHD2 appeared to have lost the ability to regulate F1FO-ATPase assembly.In addition,mass spectrometry and immunoprecipitation showed that there was an interaction between CHCHD2 and F1FO-ATPase.Three weeks after transfection with AAV-CHCHD2 T61I,we intraperitoneally injected 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine into mice to establish an animal model of chronic Parkinson's disease and found that exogenous expression of the mutant protein worsened the behavioral deficits and dopaminergic neurodegeneration seen in this model.These findings suggest that WT CHCHD2 can alleviate mitochondrial dysfunction in PD by maintaining F1F0-ATPase structure and function.

Molecular Cloning and Expression of Carotenogenic Genes in Yellowish and Mutant Whitish Loquat (Eriobotrya japonica) Fruits

ObjectiveThe study aimed to explore the factors regulating carotenoid accumulation in flesh color. MethodA loquat mutation （red-or orange-fleshed plant emerged a bud mutation of white-flesh in trunk） was used as material; HPLC analysis of β-carotene content was conducted. ResultThe β-carotene concentration in the flesh of wild and mutant types was 60.9 and 4.6 μg/g fresh weight, respectively. According to the conserved regions of genes from rose family genome, carotenogenic gene fragments in wild and mutant types were obtained. No nucleotide variation of the carotenogenic gene fragments was observed between wild and mutant genome. Real-time quantitative polymerase chain reaction （Q-PCR） was compared and one carotenogenic gene, β-ring hydroxylase （HYB） were considerably suppressed in mature mutant loquat fruits compared with that in wild. The other six carotenogenic genes were also expressed but the expression patterns appeared to be not correlated with the amount of β-carotene concentration in wild loquat flesh. ConclusionThe mutant whitish loquat lacks the ability to synthesize β-carotene because of the transcriptional down-regulation of carotenogenic gene HYB.

Cloning of cytochrome P-450 2C9 cDNA from human liver and its expression in CHL cells

AIM: Using bacterial, yeast, or mammalian cell expressing a human drug metabolism enzyme would seem good way to study drug metabolism-related problems. Human cytochrome P-450 2C9(CYP2C9) is a polymorphic enzyme responsible for the metabolism of a large number of clinically important drugs. It ranks among the most important drug metabolizing enzymes in humans. In order to provide a sufficient amount of the enzyme for drug metabolic research, the CYP2C9 cDNA was cloned and expressed stably in CHL cells. METHODS: After extraction of total RNA from human liver tissue, the human CYP2C9 cDNA was amplified with reverse transcription-polymerase chain reaction (RT-PCR), and cloned into cloning vector pGEM-T. The cDNA fragment was identified by DNA sequencing and subcloned into a mammalian expression vector pREP9. A transgenic cell line was established by transfecting the recombinant vector of pREP9-CYP2C9 into CHL cells. The enzyme activity of CYP2C9 catalyzing oxidation of tolbutamide to hydroxy tolbutamide in S9 fraction of the cell was determined by high performance liquid chromatography(HPLC). RESULTS: The amino acid sequence predicted from the cDNA segment was identical to that of CYP2C9*1, the wild type CYP2C9. However, there were two base differences, i.e. 21T】C, 1146C】T, but the encoding amino acid sequence was the same, L7, P382. The S9 fraction of the established cell line metabolizes tolbutamide to hydroxy tolbutamide; tolbutamide hydroxylase activity was found to be 0.465 +/- 0.109 micromol.min(-1).g(-1) S9 protein or 8.62 +/- 2.02mol.min(-1).mol(-1) CYP, but was undetectable in parental CHL cell. CONCLUSION: The cDNA of human CYP2C9 was successfully cloned and a cell line of CHL- CYP2C9, efficiently expressing the protein of CYP2C9, was established.

成年SD大鼠嗅球中几种蛋白的免疫组化分布研究

对成年SD大鼠嗅球中tyrosinehydroxylase、calbindinD-28K、calretinin、parvalbumin及olfactorymarkerprotein几种蛋白进行了免疫组化分布的研究.结果表明:tyrosinehydroxylase主要出现在嗅球的小球层,在外网织层有很少的阳性细胞;calbindinD-28K主要出现在小球层,在外网织层和内网织层很少;calretinin在小球层、外网织层、僧帽细胞层、内网织层、粒细胞层都有分布,嗅神经层无阳性;parvalbumin在外网织层最明显,僧帽细胞层中有少量,在小球层很少;olfactorymarkerprotein只出现在嗅神经层和小球层.此研究结果与前人的相关研究结果基本接近,但也存在一些差异.

Neuroprotective Effects of Modafinil on MPTP Mouse Model of Parkinson′s Disease

Aim To observe the neuroprotective effects of modafinil on the Parkinson'sdisease ( PD ) model induced by 1-methyl-4-phenyl-1, 2,3, 6-tetrahydropyridine (MPTP ). Methods Themodel of PD was induced by intraperitoneal injection of MPTP into C57BL/6J mice for 4 d. Modafinil(ip, 50 or 100 mg·kg^(-1)·d^(-1)) was administered following MPTP for 4 d and for another 10 dconsecatirely. The effects of modafinil on the locomotor activity, and the incubation, maintenanceperiod and grade of the tremor, the duration of the climbing rod of mouse, and the distribution ofpositive cells of ty-rosine hydroxylase (TH) and Nissl bodies in the striatum and substantia nigra(SN) were observed. The contents of dopam-ine (DA) , noradrenaline (NA) and 5-hydroxytryptamine(5-HT) in the striatum were determined. Results Modafinil (50 and 100 mg·kg^(-1)) significantlyprevented the locomotor, the tremor and climbing rod defect behavior in a dose-dependent manner (P <0.05 and P < 0.01, n = 10), prevented the decrease in the number of TH-positive cells and Nisslbodies (P<0.05, n=10), and reduced the decrease of DA, NA, and 5-HT in the striatum (P < 0.05, n =10) induced by MPTP. Conclusion Modafinil improves the behavioral deficits and prevents themonoaminergic neuron lesion in seriously impaired MPTP mouse model.

基于保护剂筛选及优化策略提高苯丙氨酸羟化酶热稳定性

苯丙氨酸羟化酶(Phenylalanine hydroxylase,PAH)具有治疗苯丙酮尿症(phenylketonuria,PKU)的潜在药用价值,而热稳定性和储存稳定性差是限制其应用的重要因素。文中考察了多种保护剂对PAH热稳定性的影响,表明添加10%的甘油可将PAH在50℃处理10 min的酶活保留率提高3.1倍,达到(98.3±0.8)%,同时海藻糖、棉籽糖和甘露醇均能将PAH酶活保留率提高到78%以上。鉴于甘油在临床应用过程中可能出现不良反应,对后3种保护剂进行正交实验,获得最佳复合保护剂组合:2.5 mmol/L甘露醇、1 mmol/L棉籽糖和1.5 mmol/L海藻糖,可将50℃酶活保留率提高3.1倍,达到(99.3±1.2)%。10%甘油和复合保护剂均能显著提高PAH在4、20、37℃的储存稳定性,其中添加10%甘油或复合保护剂使得PAH在37℃下储存半衰期分别延长到129.9 h和237.5 h,比未添加保护剂的分别提高了1.9倍和4.3倍。该结果为PAH在医药领域的应用奠定了基础。

Effect of α-synuclein on the promoter activity of tyrosine hydroxylase gene

Objective To approach the associated mechanism by which α-synuclein （α-Syn） might regulate the metabolism of dopamine. Methods A DNA fragment, located at --495 to ＋25 of the human tyrosine hydroxylase （TH） gene, was amplified by PCR and inserted into the pGL3-Basic luciferase reporter vector. The recombinant plasmid pGL3-THprom was transfected into a dopammergic cell line MES23.5 or a α-Syn over-expressed MES23.5 （named MES23.5/hα-Syn^＋）. The promoter activity was detected by the Dual Luciferase Assay System. Results The luciferase activities in the MES23.5 cells transfected with pGl.,3-Basic, pGL3-THprom, and pGL3-Control vectors were 5.60±0.67, 26.80±4.11, and 32.90±4.75, respectively. On the other hand, the luciferase activity of pGL3-THprom in the MES23.5 （26.80±4.11） was significantly higher than that in the MES23.5/hα-Syn^＋（14.40±0.61） （P〈0.01）. Conclusion These results indicate that the -495 to ＋25 region in the TH gene possesses promoter activity for controlling the gene expression, and that α-Syn may negatively regulate the metabolism of dopamine by affecting the function of TH promoter as a trans-acting factor.

Effects of Diazepam,Phenobarbital,Propranolol,and Cimetidine on Diazepam Oxidizing Isoenzymes in Rat Liver Microsomes

Isolation and identification of the liver microsomal cytochrome P 450 isoen zymes responsible for the formation of diazepam main metabolites nordiazepam and temazepam in rats were studied. The effects of P 450 inducers and inhibitors on the protein contents in SDS poly acrylamide gel electrophoresis and thin layer chromatography to the corresponding diazepam me tabolizing activities of rat liver microsomes were observed. The P 450 contents were dramatically re duced by ip diazepam, cimetidine or propranolol. Diazepam and propranolol inhibited temazepam formation, high dose of propranolol also inhibited nordiazepam formation. Phenobarbital increased the P 450 contents and induced the production of both nordiazepam and temazepam. It also induced proteins with molecular weight (m) of 51 and 59 kDa in SDS PAGE and those with m ranging from 45 to 55 kDa and from 55 to 65 kDa in TLC. Propranolol inhibited both fractions, especially that of m 55～65 kDa, whereas diazepam tended to inhibit the fraction of 45～55 kDa. The protein of m 51 kDa could be mainly involved in diazepam C3 hydroxylation, whereas those of m 59 kDa could be responsible for the N demethylation of diazepam in rats.

非典型21-羟化酶缺陷症研究进展

先天性肾上腺皮质增生症（congennital adrenal hyperplasia,CAH）是由于肾上腺皮质激素合成过程中所需的酶缺乏所致的一组常染色体隐性遗传性疾病。21-羟化酶缺乏症（21-hydroxylase deficiency,21-OHD）是CAH最常见的一种,约占90%~95%。根据临床表现的严重程度分为典型类型和非典型类型两种。

21-羟化酶缺乏症患儿的生长发育及治疗

先天性肾上腺皮质增生症（congenital adrenal hyperplasia,CAH）是一组常染色体隐性遗传疾病,由肾上腺皮质激素合成途径中酶的缺陷所引起。最常见的酶缺陷类型为21-羟化酶缺乏（21-hydroxylase deficiency,21-OHD）,占酶缺陷类型的90%,根据酶的缺陷程度又分为经典型及非经典型两种临床类型,儿童中主要为经典型。

Ginsenoside Rb1 protects dopaminergic neurons from inflammatory injury induced by intranigral lipopolysaccharide injection

Accumulating studies suggest that neuroinflammation characterized by microglial overactivation plays a pivotal role in the pathogenesis of Parkinson’s disease.As such,inhibition of microglial overactivation might be a promising treatment strategy to delay the onset or slow the progression of Parkinson’s disease.Ginsenoside Rbl,the most active ingredient of ginseng,reportedly exerts neuroprotective effects by suppressing inflammation in vitro.The present study aimed to evaluate the neuroprotective and anti-inflammatory effects of ginsenoside Rbl in a lipopolysaccharide-induced rat Parkinson’s disease model.Rats were divided into four groups.In the control group,sham-operated rats were intraperitoneally administered normal saline for 14 consecutive days.In the ginsenoside Rbl group,ginsenoside Rb1(20 mg/kg)was intraperitoneally injected for 14 consecutive days after sham surgery.In the lipopolysaccharide group,a single dose of lipopolysaccharide was unilaterally microinjected into the rat substantial nigra to establish the Parkinson’s disease model.Lipopolysaccharide-injected rats were treated with normal saline for 14 consecutive days.In the ginsenoside Rbl +lipopolysaccharide group,lipopolysaccharide was unilaterally microinjected into the rat substantial nigra.Subsequently,ginsenoside Rbl was intraperitoneally injected for 14 consecutive days.To investigate the therapeutic effects of ginsenoside Rbl,behavioral tests were performed on day 15 after lipopolysaccharide injection.We found that ginsenoside Rbl treatment remarkably reduced apomorphine-induced rotations in lipopolysaccharide-treated rats compared with the lipopolysaccharide group.To investigate the neurotoxicity of lipopolysaccharide and potential protective effect of ginsenoside Rbl,contents of dopamine and its metabolites in the striatum were measured by high-performance liquid chromatography.Compared with the lipopolysaccharide group,ginsenoside Rbl obviously attenuated the lipopolysaccharide-induced depletion of dopamine and its metabolites in the striatum.To further explore the neuroprotective effect of ginsenoside Rbl against lipopolysaccharide-induced neurotoxicity,immunohistochemistry and western blot assay of tyrosine hydroxylase were performed to evaluate dopaminergic neuron degeneration in the substantial nigra par compacta.The results showed that lipopolysaccharide injection caused a large loss of tyrosine hydroxylase-immunoreactive neurons in the substantia nigra and a significant decrease in overall tyrosine hydroxylase expression.However,ginsenoside Rb1 noticeably reversed these changes.To investigate whether the neuroprotective effect of ginsenoside Rbl was associated with inhibition of lipopolysaccharide-induced microglial activation,we examined expression of the microglia marker Iba-1.Our results confirmed that lipopolysaccharide injection induced a significant increase in Iba-1 expression in the substantia nigra;however,ginsenoside Rbl effectively suppressed lipopolysaccharide-induced microglial overactivation.To elucidate the inhibitory mechanism of ginsenoside Rb1,we examined expression levels of inflammatory mediators(tumor necrosis factor-a,interleukin-1β,inducible nitric oxide synthase,and cyclooxygenase 2)and phosphorylation of nuclear factor kappa B signaling-related proteins(IκB,IKK)in the substantia nigra with enzyme-linked immunosorbent and western blot assays.Our results revealed that compared with the control group,phosphorylation and expression of inflammatory mediators IκB and IKK in the substantia nigra of lipopolysaccharide group rats were significantly increased;whereas,ginsenoside Rbl obviously reduced lipopolysaccharide-induced changes on the lesioned side of the substantial nigra par compacta.These findings confirm that ginsenoside Rbl can inhibit inflammation induced by lipopolysaccharide injection into the substantia nigra and protect dopaminergic neurons,which may be related to its inhibition of the nuclear factor kappa B signaling pathway.This study was approved by the Experimental Animal Ethics Committee of Shandong University of China in April 2016(approval No.KYLL-2016-0148).

Rapamycin reverses ferroptosis by increasing autophagy in MPTP/MPP+-induced models of Parkinson's disease

Parkinson’s disease is a neurodegenerative disorder,and fe rroptosis plays a significant role in the pathological mechanism underlying Parkinson’s disease.Rapamycin,an autophagy inducer,has been shown to have neuroprotective effects in Parkinson’s disease.However,the link between rapamycin and ferroptosis in Parkinson’s disease is not entirely clear.In this study,rapamycin was administe red to a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinson’s disease mouse model and a 1-methyl-4-phenylpyridinium-induced Parkinson’s disease PC12 cell model.The results showed that rapamycin improved the behavioral symptoms of Parkinson’s disease model mice,reduced the loss of dopamine neurons in the substantia nigra pars compacta,and reduced the expression of ferroptosis-related indicators(glutathione peroxidase 4,recombinant solute carrier family 7,member 11,glutathione,malondialdehyde,and reactive oxygen species).In the Parkinson’s disease cell model,rapamycin improved cell viability and reduced ferro ptosis.The neuroprotective effect of rapamycin was attenuated by a ferroptosis inducer(methyl(1S,3R)-2-(2-chloroacetyl)-1-(4-methoxycarbonylphenyl)-1,3,4,9-tetrahyyridoindole-3-carboxylate)and an autophagy inhibitor(3-methyladenine).Inhibiting ferro ptosis by activating autophagy may be an important mechanism by which rapamycin exerts its neuroprotective effects.Therefo re,the regulation of ferroptosis and autophagy may provide a therapeutic target for drug treatments in Parkinson’s disease.

Laterodorsal Tegmentum and Pedunculopontine Tegmental Nucleus Circuits Regulate Renal Functions: Neuroanatomical Evidence in Mice Models

Neurons in the laterodorsal tegmentum (LDTg) and pedunculopontine tegmental nucleus (PPTg) play important roles in central autonomic circuits of the kidney. In this study, we used a combination of retrograde tracers pseudorabies virus (PRV)-614 and fluorescence immunohistochemistry to characterize the neuroanatomic substrate of PPTg and LDTg innervating the kidney in the mouse. PRV-614-infected neurons were retrogradely labeled in the rostral and middle parts of LDTg, and the middle and caudal parts of PPTg after tracer injection in the kidney. PRV-614/TPH double-labeled neurons were mainly localized in the rostral of LDTg, whereas PRV-614/TH neurons were scattered within the three parts of LDTg. PRV-614/TPH and PRV-614/TH neurons were located predominantly in the caudal of PPTg (cPPTg). These data provided direct neuroanatomical foundation for the identification of serotonergic and catecholaminergic projections from the mid-brain tegmentum to the kidney.

Treadmill step training promotes spinal cord neural plasticity after incomplete spinal cord injury

A large body of evidence shows that spinal circuits are significantly affected by training, and that intrinsic circuits that drive locomotor tasks are located in lumbosacral spinal segments in rats with complete spinal cord transection. However, after incomplete lesions, the effect of treadmil training has been debated, which is likely because of the difficulty of separating spontaneous stepping from specific training-induced effects. In this study, rats with moderate spinal cord contusion were sub-jected to either step training on a treadmil or used in the model （control） group. The treadmil training began at day 7 post-injury and lasted 20 ± 10 minutes per day, 5 days per week for 10 weeks. The speed of the treadmil was set to 3 m/min and was increased on a daily basis according to the tolerance of each rat. After 3 weeks of step training, the step training group exhibited a sig-nificantly greater improvement in the Basso, Beattie and Bresnahan score than the model group. The expression of growth-associated protein-43 in the spinal cord lesion site and the number of tyrosine hydroxylase-positive ventral neurons in the second lumbar spinal segment were greater in the step training group than in the model group at 11 weeks post-injury, while the levels of brain-derived neurotrophic factor protein in the spinal cord lesion site showed no difference between the two groups. These results suggest that treadmil training significantly improves functional re-covery and neural plasticity after incomplete spinal cord injury.

Changes of 5-hydroxytryptamine and tryptophan hydroxylase expression in the ventral horn of spinal cord

Objective To investigate changes of 5-hydroxytryptamine （5-HT） and its synthesis rate-limiting enzyme tryp-tophan hydroxylase （TPH） in the ventral horn of spinal cord after exercise-induced fatigue, and to further discuss the mecha- nism of exercise-induced central fatigue at spinal level. Methods Sixteen healthy adult Wistar rats were randomly divided into 2 groups： exercise-induced fatigue group and control group. Immunohistochemical staining for 5-HT and TPH in the ventral horn were performed and analysized quantitatively. The mean optic densities of 5-HT and TPH positive fibers or terminals were measured by computerized image analyzer. Results Both 5-HT and TPH positive fibers/terminals decreased in the exercise-induced fatigue group. The immunohistochemical staining was weaker and the mean optic densities decreased obviously in the fatigue group compared with those in the control group （P 〈 0.05）. Conclusion 5-HT and TPH in the ventral horn of spinal cord might be involved in exercise-induced fatigue.

Bone marrow-derived mesenchymal stem cells increase dopamine synthesis in the injured striatum

Previous studies showed that tyrosine hydroxylase or neurturin gene-modified cells transplanted into rats with Parkinson＇s disease significantly improved behavior and increased striatal dopamine content. In the present study, we transplanted tyrosine hydroxylase and neurturin gene-modified bone marrow-derived mesenchymal stem cells into the damaged striatum of Parkinson＇s disease model rats. Several weeks after cell transplantation, in addition to an improvement of motor function tyrosine hydroxylase and neurturin proteins were up-regulated in the injured striatum, and importantly, levels of dopamine and its metabolite 3,4-dihydroxyphenylacetic acid increased significantly. Furthermore, the density of the D2 dopamine receptor in the postsynaptic membranes of dopaminergic neurons was decreased. These results indicate that transplantation of tyrosine hydroxylase and neurturin gene-modified bone marrow-derived mesenchymal stem cells increases dopamine synthesis and significantly improves the behavior of rats with Parkinson＇s disease.

In vitro Effect of Dithiocarbamate Pesticides and of CaNa_2EDTA on Human Serum Dopamine-β-hydroxylase

Serum dopamine-β-hydroxylase (DBH) inhibition has been reported in lead workers treated with CaNa2EDTA and in alcoholic patients repeatedly treated with the alcohol aversive drug Disulfiram. The mechanism of inhibition involves Cu-chelation at the active site of DBH. The effect of CaNa2EDTA and Disulfiram on serum DBH has been compared to the effect of dithiocarbamate pesticides in vitro for the possible use of serum DBH determination for the biological monitoring of workers exposed to these pesticides.Most dithiocarbamates inhibit human serum DBH at micromoIar concentrations (range of I50, 0.027-1.6 μmol/L). The inhibitory potency increased from methyl- and dimethyl dithiocarbamates to diethyl dithiocarbamates up to the most potent ethylene bisdithiocarbamates. The I50 of CaNa.EDTA was 3.8 mol/L, higher than those of dithiocarbamates. Copper addition to the test system rcactivated at stoichiometric concentrations dithiocarbamaie-inhibited DBH indicating that both base line values and percent of inhibition can be calculated in a single blood sample. Results suggest that serum DBH determination couId be useful in case of acute poisoning involving high doses of dithiocarbamate pesticides

Downregulation of thioredoxin reductase 1 expression in the substantia nigra pars compacta of Parkinson's disease mice

Because neurons are susceptible to oxidative damage and thioredoxin reductase 1 is extensively distributed in the central nervous system and has antioxidant properties, we speculated that the enzyme may be involved in the pathogenesis of Parkinson＇s disease. A Parkinson＇s disease model was produced by intraperitoneal injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine into C57BL/6 mice. Real-time reverse transcription-PCR, western blot analysis and colorimetric assay showed that the levels of thioredoxin reductase 1 mRNA and protein were decreased, along with a significant reduction in thioredoxin reductase activity, in the midbrain of Parkinson＇s disease mice compared with normal mice. Immunohistochemical staining revealed that the number of thioredoxin reductase 1-positive neurons in the substantia nigra pars compacta of Parkinson＇s disease mice was significantly decreased compared with normal mice. These experimental findings suggest that the expression of thioredoxin reductase 1 in the substantia nigra pars compacta of Parkinson＇s disease mice is significantly decreased, and that the enzyme may be associated with disease onset.

Changes of microbial community structures and functional genes during biodegradation of phenolic compounds under high salt condition

The changes of microbial community structures and functional genes during the biodegradation of single phenol and phenol plus p-cresol under high salt condition were explored.It was found that the phenol-fed system（PFS） exhibited stronger degrading abilities and more stable biomass than that of the phenol plus p-cresol-fed system（PCFS）.The microbial community structures were revealed by a modern DNA fingerprint technique,ribosomal intergenic spacer analysis（RISA）.The results indicated that the microbial community of PFS changed obviously when gradually increased phenol concentration,while PCFS showed a little change.16S rRNA sequence analysis of the major bands showed that Alcanivorax sp.genus was predominant species during phenolic compounds degradation.Furthermore,amplified functional DNA restriction analysis（AFDRA） on phenol hydroxylase genes showed that the fingerprints were substantially different in the two systems,and the fingerprints were not the same during the different operational periods.