Novel histochemical approaches to the prealbumin-related senile and familial forms of systemic amyloidosis

The immunoperoxidase method, the autoclave method, and a newly developed alkaline-guanidine method were used to distinguish senile (SSA) and familial types (FAP) of prealbumin-related amyloidosis in formalin-fixed, paraffin-embedded tissue sections. Because all the amyloid deposits of SSA and FAP reacted positively with the antiprealbumin antiserum, a classification of the amyloid fibril proteins of FAP and SSA by immunohistochemistry, using polyclonal anti-prealbumin antisera, was not feasible. Both the senile and familial forms of amyloidosis showed unchanged Congophilia after prolonged autoclaving. In the alkaline-guanidine method, FAP amyloids were resistant to incubation for 2 hours. On the other hand, amyloid deposits of SSA lost the Congophilia and green birefringence with 2 hours' alkaline-guanidine treatment. Therefore, the autoclave method combined with the alkaline-guanidine method will considerably facilitate differentiation of SSA and FAP, without specific antisera.     

Immunohistochemical confirmation of Creutzfeldt-Jakob disease with a long clinical course with amyloid plaque core antibodies.

Amyloid plaques have been found in the brains of some patients with Creutzfeldt-Jakob disease (CJD) and all patients with Gerstmann-Sträussler syndrome (GSS). We examined paraffin sections from 45 patients with CJD or GSS and from 51 patients with other neurologic diseases, using an antiserum against GSS amyloid plaque cores. The GSS amyloid plaque core antiserum revealed not only birefringent amyloid plaques but also small plaques that cannot be detected by the staining with Congo red dye. Positive immunolabeling was demonstrated in 59% of 34 Japanese patients with CJD, in 100% of 11 patients with GSS, and in none with other neurologic diseases. All cases of CJD of short duration (less than 11 months) were evaluated as being negative, and 95% of 21 long survivors (over 12 months) were positive. This immunohistochemical approach revealed that amyloid plaque is a hallmark of CJD with a long clinical course.     

Efficacy of early use of continuous isoproterenol inhalation therapy for severe asthma attacks in children]

The aim of this study was to evaluate the efficacy and safety of continuous isoproterenol inhalation therapy for asthma attacks in children. We used l-body isoproterenol (Proternol L) in 22 children with 32 episodes of severe attacks. One of them did not respond to this therapy, and two had complications (atelectasis and pneumothorax). Twenty-nine cases were divided into three subgroups according to their clinical scores; A) scores less than or equal to 4, which meant that they were in the early stage of severe attack (n = 9), B) scores 5-6, which meant impending respiratory failure (n = 17), C) scores greater than or equal to 7, which meant respiratory failure (n = 3). The values of SpO2 at the start of this therapy were 94.8, 91.5, 82.0%, respectively. The more severe their attacks were, the lower their SpO2 levels were. The periods until their scores became zero were 0.78, 6.3, 17.2 hours, respectively. There were significant differences between each period respectively (p less than 0.001, p less than 0.01). Heart rates decreased when their symptoms improved, and other adverse effects were not detected. These results suggest that this therapy is effective and safe for children with severe asthma attacks, especially in the early stage.     

Incidence and characterization of age related amyloid deposits in the human anterior pituitary gland

Summary To identify amyloid deposits in the anterior pituitary gland, we have immunohistochemical, histochemical and alkaline Congo red staining. The anti-human P component reacted positively with these amyloid deposits, while antisera against prealbumin, AA type amyloid fibril protein and various anterior pituitary hormones were negative. A combination of Congo red and anti-human P component staining was most sensitive and reliable for detection of amyloid in the anterior pituitary glands of 300 randomly autopsied patients. Amyloid deposits increased in parallel with the age of the patients, however, they appeared earlier and more frequently than heretofore reported. Deposition of amyloid was seen initially in the 3rd decade and the positivity rate of amyloid deposits was 73% in the 5th decade. The histochemical characteristics of these pituitary amyloid deposits differed from those of cerebral and systemic deposits, particularly those found in the amyloid of senile systemic amyloidosis.This study was supported in part by a grant from the Fundation for Advancement of Clinical Medicine and Ministry of Health and Welfare of Japan     

Neural crest origin of clear cell sarcoma of tendons and aponeuroses

Summary In order to clarify the histogenesis of clear cell sarcoma of tendons and aponeuroses (CCS), two cases of human and one nude mouse-transplanted CCS line were studied using an ultrastructural and enzyme cytochemical approach. Most of the tumour cells obtained from the primary and transplanted CCS demonstrated melanosomes in various stages of development within the cytoplasm, whereas no melanosomes could be identified in the metastatic CCS. However, cholinesterase and tyrosinase activities could be demonstrated not only in the melanotic primary and transplanted CCS but also in the amelanotic metastatic CCS. The results therefore support the hypothesis that CCS is a soft tissue tumour derived from the neural crest.     

Two dinucleotide repeat polymorphisms at the D8S1218 and D8S1219 loci

Summary Two polymorphic dinucleotide (CA) repeat dones were isolated from a CEPH mega-YAC clone (844E2), and were localized to chromosome 8 using a panel of 13 mouse/human somatic cell hybrids.     

Negative chronotropic actions of endothelin-1 on rabbit sinoatrial node pacemaker cells

1. The effects of endothelin-1 (ET-1) on sinoatrial (SA) node preparations of the rabbit heart were studied by means of whole-cell clamp techniques.
2. ET-1 at 1 nM slowed the spontaneous beating activity and rendered half of the cells quiescent. At a higher concentration of 10 nM, the slowing and cessation of spontaneous activity were accompanied by hyperpolarization.
3. In voltage-clamp experiments, ET-1 decreased the basal L-type Ca²⁺ current (I_Ca(L)) dose-dependently with a half-maximal inhibitory concentration (EC₅₀) of 0.42 nM and maximal inhibitory response (E_max) of 49.5%. The delayed rectifying K⁺ current (I_K) was also reduced by 33.2±11.1% at 1 nM. In addition, an inwardly rectifying K⁺ current was activated by ET-1 at higher concentrations (EC₅₀=4.8 nM). These ET-1-induced changes in membrane currents were abolished by BQ485 (0.3 μM), a highly selective ET_A receptor antagonist.
4. When I_Ca(L) was inhibited by ET-1 (1 nM), subsequent application of 10 μM ACh showed no additional decrease in I_Ca(L), suggesting the involvement of cyclic AMP in the effects of ET-1 on I_Ca(L). In contrast, 1 nM ET-1 further decreased I_Ca(L) in the presence of 10 μM ACh, suggesting that ET-1 activates some additional mechanism(s) which inhibit I_Ca(L). The ET-1-induced I_Ca(L) inhibition was abolished by protein kinase A inhibitory peptide (PKI, 20 μM) or H-89 (5 μM). However, the I_Ca(L) inhibition was not affected by methylene blue (10 μM), suggesting a minor role for cyclic GMP in the effect of ET-1 under basal conditions.
5. ET-1 failed to inhibit I_Ca(L) when the pipette contained GDPβS (200 μM). However, incubation of the cells with pertussis toxin (PTX, 5 μg ml⁻¹, >6 h) only reduced the ET-1-induced inhibition to 21.5±9.5%, whereas it abolished the inhibitory effect of ACh on I_Ca(L).
6. Intracellular perfusion of 8-bromo cyclicAMP (8-Br cyclicAMP, 500 μM) attenuated, but did not abolish the inhibitory effect of ET-1 on I_Ca(L). This 8-Br cyclicAMP-resistant component (17.5±14.4%, n=20) was not affected by combined application of 8-Br cyclicAMP with 8-bromo cyclicGMP (500 μM), ryanodine (1 μM) or phorbol-12-myristate-13-acetate (TPA; 50 nM).
7. In summary, ET-1 exerts negative chronotropic effects on the SA node via ET_A-receptors. ET-1 inhibits both I_Ca(L) and I_K, and increases background K⁺ current. The inhibition of I_Ca(L) by ET-1 is mainly due to reduction of the cyclicAMP levels via PTX-sensitive G protein, but some other mechanism(s) also seems to be operative.

     

PCNA immunostaining combined with AgNOR staining in esophageal squamous cell carcinoma to identify patients with a poor prognosis

Immunostaining of the proliferating cell nuclear antigen (PCNA) provides important information about cell kinetics and is easily performed on routinely obtained formalin-fixed, paraffin-embedded materials. We report herein the results of a retrospective study on PCNA staining in esophageal cancer undertaken to determine its significance. As this study indicated that immunoreactivity was preserved in specimens fixed within 24 h, only 31 specimens from surgical patients were available for this investigation. The mean PCNA index of the patients without invasion to the adventitia (35.7±17.9) was significantly lower than that of those with invasion to the adventitia or neighboring structures (49.7±14.5), while the PCNA index did not correlate with other clinicopathologic parameters such as histologic type, lymph node metastases, or prognosis. However, when an analysis of PCNA staining was combined with an analysis of argyrophilic nucleolar organizer region (AgNOR) staining, a correlation with prognosis was found. In fact, seven patients with a high PCNA index (44) and AgNOR count (6) had a significantly poorer prognosis than the remaining 22 (P=0.0014), and six of these seven patients died within 2 years. These results indicate that this combined evaluation may be useful for the identification of patients with a poor prognosis among those undergoing surgery for esophageal squamous cell carcinoma.     

Hypoalbuminemia accelerates erythrocyte membrane lipid peroxidation in chronic hemodialysis patients

BACKGROUND: Persistent hypoalbuminemia is a long-term poor prognostic factor in chronic hemodialysis patients. PATIENTS AND METHODS: We investigated the correlation between the degree of peroxidation of erythrocyte membrane lipids, erythrocyte alpha tocopherol content, erythrocyte glutathione peroxidase activity and serum albumin concentration in twelve patients with uremia not undergoing hemodialysis and fifteen patients on maintenance hemodialysis. RESULTS: The glutathione peroxidase activity in erythrocytes was higher in patients of uremia not undergoing hemodialysis than in chronic hemodialysis patients. A significant negative correlation was observed between the erythrocyte alpha tocopherol content and the degree of erythrocyte membrane lipid peroxidation in chronic hemodialysis patients. There was a statistically significant difference in the degree of erythrocyte membrane lipid peroxidation between patients with chronic hemodialysis-associated hypoalbuminemia and chronic hemodialysis patients having normal serum albumin levels. CONCLUSION: This study suggested that serum albumin inhibits peroxidation of erythrocyte membrane lipids and that hemodialysis induces recovery of serum reductivity. We conclude that persistent hypoalbuminemia worsens the serum antioxidant activity in chronic hemodialysis patients and may contribute to increased oxidative cell damage.