Corneal crystals in nephropathic cystinosis: natural history and treatment with cysteamine eyedrops

Although renal disease is the most prominent feature of the lysosomal storage disease cystinosis, corneal cystine crystal formation remains a major complication, leading to photophobia, corneal erosions, and keratopathies. Moreover, the extent of corneal crystal accumulation reflects the course and severity of the disease itself, and the cornea is accessible to direct examination. Therefore, we employed a scoring system, based on a library of slit-lamp photographs of corneas with increasing crystal densities (0.00-3.00), to assess the degree of crystal accumulation in 170 patients with nephropathic cystinosis examined at the National Institutes of Health between 1976 and 2000. None of the patients had received topical cystine-depleting therapy at the time of the evaluation. In this natural history study, infants in the first year of life had absent or minimal corneal crystals, i.e., a corneal cystine crystal score (CCCS) of 0 or 0.25. However, the CCCS increased linearly with age, such that every patient had visible crystals by 16 months of age, and plateaued at approximately 3.00 by early adolescence. Longitudinal studies in representative patients support the cross-sectional results. Individuals homozygous for the common 57-kb deletion involving the cystinosis gene (CTNS) displayed the same course of corneal crystal accumulation as did individuals not bearing the large deletion. Patients with ocular or nonnephropathic cystinosis had CCCSs that were, in general, half those expected for patients with nephropathic cystinosis of the same age. Administration of 0.55% cysteamine eyedrops, given 6 to 12 times per day, dissolved corneal cystine crystals in 10 representative patients with nephropathic cystinosis aged 1 to 32 years within 8 to 41 months.     

Analysis of the CTNS gene in patients of German and Swiss origin with nephropathic cystinosis

The autosomal recessive lysosomal storage disorder, nephropathic cystinosis is characterized by impaired transport of free cystine out of lysosomes. The gene responsible for cystinosis, CTNS, consists of 12 exons and encodes a 55 kDa putative lysosomal membrane protein, called cystinosin. Up to now more than 55 different CTNS mutations have been described in cystinosis. We have analyzed the mutation pattern in a population of 40 cystinosis patients from 35 families of German and Swiss origin. CTNS mutations in 68 out of 70 alleles were identified. The common 57-kb deletion accounted for 65% of the alleles. In five patients we found a known GACT deletion at position 18-21. In two patients we identified a nucleotide substitution at codon 339 and one patient showed a CG insertion at position 697-698. In five patients we observed a G insertion at position 926-927. Moreover, five novel mutations including two deletions involving exon 3 (61-61+2delGGT) and exon 6 (280delG), two insertions in exon 6 (292-293insA) and exon 7 (684insCACTT) and one nucleotide substitution in exon 11 (923G>T) have been identified. These data provide a basis for routine molecular diagnosis of cystinosis in the central European population, especially in cystinosis patients of German and Swiss origin.     

Quantitative,functional, morphological and ultrastructural recovery of platelets as predictor for cryopreservation

Cryopreservation of platelets is of great interest, since it could extend the shelf life of therapeutic platelet concentrates and facilitate stockpiling and inventory control in blood banking. Despite the use of many cryopreservation procedures the optimal cryopreservation procedure is not defined yet. We have compared the cryopreservation of human platelets by various protocols employing controlled-rate and non-controlled-rate freezing procedures in combination with different concentrations of DMSO (6% and 10%) or 5% DMSO + 6% HES combination. After storage for 1 to 3 months, samples were thawed and analyzed. Measurements included cell recovery, platelet viability according to hypotonic shock response (HSR), platelet aggregation with ADP, morphological and ultrastructural properties of defrozen platelets. Our findings show that the application of our original procedure for controlled-rate freezing consisting of six cooling steps (cooling rate 1 degree C/min) with compensation of released heat of fusion (cooling rate 2 degrees C/min) has significantly influenced the quality of thawed platelets. At the same time, a concentration of 6% DMSO proved to be the most effective. In summary, cryopreservation of human platelets using controlled-rate freezing procedure in combination with lower (6%) DMSO concentration resulted in less damage from freezing and higher recovered function of platelets.     

Oncocytoma of the prostate: a case report with ultrastructural and immunohistochemical evaluation

An oncocytoma of the prostate of an 87-year-old man, who underwent transurethral resection because of prostatic enlargement, is reported. Electronmicroscopic and immunohistochemical evaluation confirmed the oncocytic nature of the tumour cells. To our knowledge, this lesion is hitherto undescribed in this location.     

Elastofibroma: a histochemical, immunohistochemical, and ultrastructural study of two patients

Elastofibroma is a rare neoplasm. In this article, we report our study of the pathogenesis of fibrosis in elastofibroma. Three tumors obtained from two patients were selected. One patient was a 57-year-old Japanese woman who had a bilateral tumor, and the other patient was a 83-year-old Japanese man. All tumors occurred in the infrascapular region. Macroscopically, the cut surface of all tumors showed a poorly defined and whitish mass with yellowish foci. Microscopically, the tumor consisted of collagen fiber bundles, abnormal elastic fibers, and spindle cells suggestive of fibroblasts. Elastica-van Gieson and Masson-trichrome stain identified abnormal elastic fibers and abundant collagen fibers, respectively, in elastofibroma. Immunohistochemically, fibroblasts were positive for CD34 but negative for alpha-smooth muscle actin and h-caldesmon. Additionally, the cytoplasm of many fibroblasts was positive for TGF-beta in all tumors. Ultrastructurally, some fibroblasts with abundant organelles in one tumor were observed in the adjacent area to amorphous elastic mass and bundles of collagen fibers. However, no myofibroblasts were ultrastructurally identified in the tumor. Finally, our study supplies further evidence that elastofibroma may show the proliferation of CD34-positive fibroblasts and contain no myofibroblasts, and that fibroblasts may produce both abnormal elastic fibers and collagen fibers through the secretion of TGF-beta.     

Histomorphometric, ultrastructural and microhardness evaluation of the osseointegration of a nanostructured titanium oxide coating by metal-organic chemical vapour deposition: an in vivo study

Over the past decade the increase of elderly population has determined a rise in the incidence of bone fractures, and the improvement of the implant-bone interface remains an open problem. Metal-organic chemical vapour deposition (MOCVD) has recently been proposed as a technique to coat orthopaedic and dental prostheses with metal nanostructured oxide films either through the decomposition of oxygenated compounds (single-source precursors) or the reaction of oxygen-free metal compounds with oxygenating agents. The present study was performed to assess the in vivo biocompatibility of commercially pure Ti (control material: TI/MA) implants ( psi 2 mm x 5 mm length) coated with nanostructured TiO2 films by MOCVD (Ti/MOCVD) and then inserted into rabbit femoral cortical (middhiaphysis) and cancellous (distal epiphysis) bone. Histomorphometric, ultrastructural and microhardness investigations were carried out. Four and 12 weeks after surgery, significant (p<0.0005) increases in AI of Ti/MOCVD implants were observed as compared to Ti/MA implants (distal femoral epiphysis: 4 weeks=8.2%, ns; 12 weeks=52.3%, p <0.005; femoral diaphysis: 4 weeks=20.2%, p <0.0005; 12 weeks=10.7%, p <0.005). Bone microhardness results showed significant increases for the Ti/MOCVD versus Ti/MA implants at 200 microm in the femoral diaphysis (4 weeks=14.2, p <0.005) and distal femoral epiphysis (12 weeks=14.5, p <0.01) at 4 and 12 weeks, respectively. In conclusion, the current findings demonstrate that the nanostructured TiO2 coating positively affects the osseointegration rate of commercially pure Ti implants and the bone mineralization at the bone-biomaterial interface in both cortical and cancellous bone.     

High magnesium concentrations in the dense bodies of human blood platelets from patients with atopic dermatitis and chronic myelogenous leukemia

Dense bodies of the normal human blood platelet contain high concentrations of phosphorus and calcium and very small amounts of magnesium, 69 mM/kg dry weight. Quantitative energy dispersive X-ray microanalysis of the platelet dense bodies on fresh air-dried smears of blood from patients with atopic dermatitis and chronic myelogenous leukemia disclosed higher concentrations of magnesium, 251 and 176 mM/kg, respectively, as well as large amounts of phosphorus and calcium. High magnesium content in the dense bodies is discussed in relation to large amounts of magnesium in the dense bodies from the blood of rats, mice and guinea pigs.     

Determination of activated platelets: evaluation of methodology and application for patients with idiopathic thrombocytopenic purpura]

Platelet activation causes a change in surface expression of several endogenous proteins, such as CD62P, CD63 and CD40L. Therefore, it is possible to analyze the functional in vivo status of the circulating platelet population directly by flow cytometry. In this study we developed the method to be suitable for use in clinical studies. We used EDTA-2K as anticoagulant since the sample anticoagulated with EDTA-2K, sodium citrate or ACD-A showed no difference in the data of activated platelets. We determined whether fixation of sample is necessary. The samples stained before or without fixation showed abnormally high level of activated platelets, indicating that fixation is necessary before staining. It is controversial whether activated platelets circulate in patients with idiopathic thrombocytopenic purpura(ITP). We measured activated platelets in patients with ITP using our optimised method. The percentages of CD62P, CD63 and CD40L positive platelets were significantly high in patients with ITP and 24%, 55% and 36% (respectively) of ITP patients showed elevated level of activated platelets. These data indicate that activated platelets circulate in ITP patients.     

The genomic region encompassing the nephropathic cystinosis gene (CTNS): complete sequencing of a 200-kb segment and discovery of a novel gene within the common cystinosis-causing deletion

Nephropathic cystinosis is an autosomal recessive disorder caused by the defective transport of cystine out of lysosomes. Recently, the causative gene (CTNS) was identified and presumed to encode an integral membrane protein called cystinosin. Many of the disease-associated mutations in CTNS are deletions, including one >55 kb in size that represents the most common cystinosis allele encountered to date. In an effort to determine the precise genomic organization of CTNS and to gain sequence-based insight about the DNA within and flanking cystinosis-associated deletions, we mapped and sequenced the region of human chromosome 17p13 encompassing CTNS. Specifically, a bacterial artificial chromosome (BAC)-based physical map spanning CTNS was constructed by sequence-tagged site (STS)-content mapping. The resulting BAC contig provided the relative order of 43 STSs. Two overlapping BACs, which together contain all of the CTNS exons as well as extensive amounts of flanking DNA, were selected and subjected to shotgun sequencing. A total of 200,237 bp of contiguous, high-accuracy sequence was generated. Analysis of the resulting data revealed a number of interesting features about this genomic region, including the long-range organization of CTNS, insight about the breakpoints and intervening DNA associated with the common cystinosis-causing deletion, and structural information about five genes neighboring CTNS (human ortholog of rat vanilloid receptor subtype 1 gene, CARKL, TIP-1, P2X5, and HUMINAE). In particular, sequence analysis detected the presence of a novel gene (CARKL) residing within the most common cystinosis-causing deletion. This gene encodes a previously unknown protein that is predicted to function as a carbohydrate kinase. Interestingly, both CTNS and CARKL are absent in nearly half of all cystinosis patients (i.e., those homozygous for the common deletion). [The sequence data described in this paper have been submitted to the GenBank data library under accession nos. AF168787 and AF163573.]     

Curcumin improves tendon healing in rats: a histological,biochemical, and functional evaluation

Curcumin, a compound extracted from the roots of Zingiberaceae, has been proposed as a treatment for tissue injury but studies are yet to be done on its effect on tendon healing. Therefore, we performed a series of experiments to test our hypothesis that curcumin has positive effects on tendon repair. Patellar tendon window defect was created in Sprague–Dawley rats and these were divided into two groups: (i) control and (ii) curcumin-treated. Curcumin (100 mg/kg body weight) was applied by oral gavage. Its potential for promoting tendon healing was assessed by histological evaluation, mRNA expression of tenocyte-related genes, malondialdehyde (MDA) levels, manganese-dependent superoxide dismutase (MnSOD) activity, quantification of hydroxyproline (HOPro), and biomechanical testing. In this tendon injury model, curcumin significantly improved the healing properties as evidenced by extensive deposition of well-organized collagen fibers, decreased MDA levels, and increase in the biomechanical properties and MnSOD activity of the regenerated tendon tissues. The current study showed that curcumin can improve the quality of tendon rupture healing, and thus represents a promising strategy in the management of injured tendon tissue.     

Protein kinase A activity in platelets from patients with bipolar disorder

BACKGROUND: Abnormal levels of protein kinase A (PKA) were found in patients with bipolar disorder (BD). Since altered levels are generally accompanied by functional modifications, the purpose of this study was to investigate PKA activity in patients with BD. METHODS: PKA activity was assessed in platelets from 20 drug-free bipolar patients and 19 controls. RESULTS: The cAMP-stimulated PKA activity was significantly increased in bipolar patients compared with controls. LIMITATIONS: This study made use of platelets, which may not fully represent changes occurring in specific brain regions. CONCLUSION: This study adds to the growing evidence suggesting that abnormalities of PKA are associated with BD.     

Time course of serotonin in platelets from patients with affective disorders

Temporal variations in platelet levels of serotonin were found to be significantly decreased in patients with endogenous affective disorders, particularly those with the bipolar type of manic-depressive psychosis. The time course of serotonin content in these cells was not affected by either the sex or the age of the patients.In vitro incubation with lithium oxybutyrate raised mean platelet serotonin levels and stabilized their fluctuation in platelets from healthy subjects but not in those from the mental patients. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 117, N ^o 3, pp. 240–242, March, 1994 Presented by E. D. Gol'dberg, Member of the Russian Academy of Medical Sciences     

Secretion by striated ducts of mammalian major salivary glands: review from an ultrastructural, functional, and evolutionary perspective

In addition to their role in electrolyte homeostasis, striated ducts (SDs) in the major salivary glands of many mammalian species engage in secretion of organic products. This phenomenon usually is manifested as the presence of small serous-like secretory granules in the apical cytoplasm of SD cells. The composition of these granules is largely unknown, except in the case of the cat and rat submandibular gland, where the granules have unequivocally been shown to contain kallikrein. In some species, the apical cytoplasm of SD cells contains variable numbers of vesicles, both spherical and elongated, that vary in appearance from 'empty' to moderately dense. In the rat parotid gland, lucent vesicles transport glycoproteins to the luminal surface where they are incorporated into the apical plasmalemma and the glycocalyx. There is a strong possibility that in various species some of these vesicles are involved in transcytosis of antibodies to the saliva from their source (plasma cells) in the surrounding connective tissue. In addition, vesicles may engage in transfer of growth factors from the saliva to the interstitium. In a few species, conventional SDs have been replaced by ducts that are wholly given over to secretion, i.e., they entirely lack basal striations; although such ducts occupy the histological position of conventional SDs, it is not clear whether they represent a new type of duct or merely are modifications of SDs. Broad-based comparisons of ultrastructural and other data about SDs offer some insight into evolutionary history of salivary glands and their role in the adaptive radiation of mammals. Evolutionary patterns emerged when we made interspecific comparisons across mammalian orders. Among the bats, there is a clear relationship between SD secretion and general categories of diet.     

Gastrointestinal stromal tumor of the stomach with rhabdoid phenotype: immunohistochemical, ultrastructural, and immunoelectron microscopic evaluation

A variety of neoplasms with rhabdoid differentiation have been reported in many sites. The authors describe a case of gastrointestinal stromal tumor (GIST) of the stomach that exhibited prominent rhabdoid features. Immunohistochemically, the tumor cells displayed positive staining for vimentin, c-kit, CD34, and alpha smooth muscle actin. Ultrastructural examination of the rhabdoid tumor cells revealed paranuclear whorls of intermediate filaments, which were immunoreactive for vimentin by both light microscopic immunohistochemical and protein A gold immunoelectron microscopic techniques. On H&E light microscopic examination alone, such a tumor could be mistaken for a variety of epithelial, mesenchymal, or other neoplasms that may show rhabdoid features. One report of GIST with a rhabdoid histologic phenotype has been described. This is the second known report of such a case with immunophenotypic and ultrastructural evaluation, and the first case with immunoelectron microscopic examination.     

Fine structural characterisation of a Rickettsia-like organism in human platelets from patients with symptoms of ehrlichiosis.

Since 1982, Ehrlichia platys infection has been diagnosed in canines from Venezuela by the use of buffy coat smears. In 1992, ehrlichia-like bodies were observed in platelets from a severely ill girl by light microscopy. The patient was seropositive to E. chaffeensis by the indirect fluorescent antibody test (IFAT). Tetracycline was administered and the patient recovered. More than 400 cases with such intra-platelet organisms have been studied at this laboratory over the past 6 years, and all the patients had a good response to the treatment. To determine whether the organisms in human blood platelets were truly platelet ehrlichiae, IFAT and transmission electron microscopy (TEM) studies were undertaken in four patients. Light microscopic examination of blood samples revealed the dense organism inside platelets, and a great reactivity of the blood cells. Sera from the four patients were seronegative against E. chaffeensis and E. platys antigens. Three of four samples contained the intra-platelet organisms when examined by TEM. Electron microscopy showed platelets with vacuoles containing pleomorphic organisms. These organisms had a thickened membrane, an electron-translucent inner area and an electron-dense granular component in the periphery. An abundant electron-dense material was observed surrounding them. The ultrastructure of such micro-organisms has not been reported previously, Based on the similarity of many of their characteristics with rickettsiae, we suggest that the microorganisms found in the present study might belong to the family Rickettsiaceae.     

Mesenteric fibromatosis: cytologic, histologic, and ultrastructural findings in a case

The cytologic findings in a case of mesenteric fibromatosis initially suggested the diagnosis of a benign spindle-cell soft-tissue tumor. Subsequent histology and electron microscopy were performed on the resected mass, and the definitive diagnosis was established. The patient had a history of previous abdominal surgery, but no features of Gardner's syndrome. The difficulties associated with the diagnosis of mesenteric fibromatosis and the cytologic diagnosis of benign and spindle-cell soft-tissue tumors and low-grade sarcomas in general are discussed.