Detection of cytokinins and auxin in plant tissues using histochemistry and immunocytochemistry.

We report a new method for histochemical localization of cytokinins (CKs) in plant tissues based on bromophenol blue/silver nitrate staining. The method was validated by immunohistochemistry using anti-trans-zeatin riboside antibody. Indole-3-acetic acid (auxin, IAA) was localized by anti-IAA antibody in plant tissues as a proof for IAA histolocalization. We used root sections, because they are major sites of CKs synthesis, and insect galls of Piptadenia gonoacantha that accumulate IAA. Immunostaining confirmed the presence of zeatin and sites of accumulation of IAA indicated by histochemistry. The colors developed by histochemical reactions in free-hand sections of plant tissues were similar to those obtained by thin layer chromatography (TLC), which reinforced the reactive sites of zeatin. The histochemical method for detecting CKs is useful for galls and roots, whereas IAA detection is more efficient for gall tissues. Therefore, galls constitute a useful model for validating histochemical techniques due to their rapid cell cycles and relatively high accumulation of plant hormones.

Epilepsy in hemiplegic migraine: Genetic mutations and clinical implications.

Objective We performed a systematic review on the comorbidities of familial/sporadic hemiplegic migraine (F/SHM) with seizure/epilepsy in patients with CACNA1A, ATP1A2 or SCN1A mutations, to identify the genotypes associated and investigate for the presence of mutational hot spots. Methods We performed a search in MEDLINE and in the Human Gene Mutation and Leiden Open Variation Databases for mutations in the CACNA1A, ATP1A2 and SCN1A genes. After having examined the clinical characteristics of the patients, we selected those having HM and seizures, febrile seizures or epilepsy. For each gene, we determined both the frequency and the positions at protein levels of these mutations, as well as the penetrance of epilepsy within families. Results Concerning F/SHM-Epilepsy1 (F/SHME1) and F/SHME2 endophenotypes, we observed a prevalent involvement of the transmembrane domains, and a strong correlation in F/SHME1 when the positively charged amino acids were involved. The penetrance of epilepsy within the families was highest for patients carrying mutation in the CACNA1A gene (60%), and lower in those having SCN1A (33.3%) and ATP1A2 (30.9%) mutations. Conclusion Among the HM cases with seizure/epilepsy, we observed mutational hot spots in the transmembrane domains of CACNA1A and ATP1A2 proteins. These findings could lead to a better understanding of the pathological mechanisms underlying migraine and epilepsy, therein guaranteeing the most appropriate therapeutic approach.

Neuropsychiatric adverse events of antiepileptic drugs in brain tumour-related epilepsy: an Italian multicentre prospective observational study.

BACKGROUND AND PURPOSE: We assessed the prevalence and magnitude of neuropsychiatric adverse events (NPAEs) associated with antiepileptic drugs (AEDs) among patients with brain tumour-related epilepsy (BTRE). METHODS: This observational, prospective, multicentre study enrolled 259 patients with BTRE after neurosurgery. All patients received AED monotherapy. Efficacy was assessed through clinical diaries, whereas NPAEs were collected using the Neuropsychiatric Inventory Test-12 questionnaire at baseline and after 5 months. RESULTS: Tumour localization in the frontal lobe was associated with a higher prevalence of NPAEs (odds ratio, 7.73; P < 0.001). Independent of tumour localization, levetiracetam (LVT) treatment was associated with higher prevalence and magnitude of NPAEs (odds ratio, 7.94; P < 0.01) compared with other AEDs. Patients with oligodendroglioma reported more NPAEs than patients with other tumour types. NPAEs were not influenced by chemotherapy, radiotherapy or steroid treatment. Evaluating non-neurobehavioural adverse events of AEDs, no significant differences were found among AEDs, although patients treated with old AEDs had a higher prevalence of adverse events than those treated with new AEDs. CONCLUSIONS: Both tumour localization in the frontal lobe and LVT treatment are associated with a higher risk of NPAEs in patients with BTRE. LVT is regarded as a first-line option in patients with BTRE because of easy titration and few significant drug-to-drug interactions. Thus, as NPAEs lead to poor compliance and a high dropout rate, clinicians need to accurately monitor NPAEs after AED prescription, especially in patients with frontal lobe tumours receiving LVT.

Carcinoma ex oncocytic Schneiderian (cylindrical cell) papilloma.

INTRODUCTION: This study reviews the clinicopathologic features of carcinoma presenting in oncocytic Schneiderian papilloma (OSP; cylindrical cell papilloma), a rare histologically distinctive papilloma sharing features with inverted papilloma. MATERIALS AND METHODS: Nine cases of carcinoma ex OSP (1970-1990), six from the files of the Armed Forces Institute of Pathology (Washington, DC) and three from the surgical pathology files, Presbyterian-University Hospital and Eye and Ear Institute (Pittsburgh, PA), were studied. One case was previously reported. Clinical data were obtained from the patients' medical records. All available histologic slides were reviewed. RESULTS: The patients (eight men, one woman) ranged in age from 48 to 83 years (mean age, 74 years). The carcinoma and OSP were synchronous in eight patients. In one case, the carcinoma occurred 8 years after OSP was diagnosed on biopsy. The presenting manifestations included nasal obstruction, "polyps," epistaxis, and proptosis. Radiographically, a sinonasal mass was present, often with extension to adjacent sites and bone destruction. Most patients received postoperative irradiation. Of six patients with follow-up, four died of carcinoma and two were alive without disease at 1.5 and 7 years, respectively. The types of carcinoma were: squamous cell (n = 6), high-grade mucoepidermoid (n = 2), and sinonasal undifferentiated (n = 1). Histologic transition was observed between carcinoma and dysplastic/benign OSP in eight cases. CONCLUSION: Demonstration of histologic continuity between carcinoma and dysplastic OSP epithelium in eight of our nine cases indicates origin of carcinoma from the OSP, rather than a coincidence of the two neoplasms. Although histologically benign, OSP should be totally excised at diagnosis because of a potential for malignant transformation. This is a US government work. There are no restrictions on its use.

Thromboxane generation and pulmonary reactivity to arachidonic acid in heart-lung preparation of guinea-pig: modulation by PCO2.

A dose-related increase of pulmonary vasoconstrictive and bronchoconstrictive effects, as well as of the amounts in the perfusing fluid of TXB2, the stable metabolite of TXA2, was obtained through administration of arachidonic acid (AA) in normocapnic and deeply hypocapnic guinea-pig heart-lung preparations (HLPs) perfused with homologous red blood cells suspended in a modified Tyrode solution. Pulmonary hypertensive effects and the amounts of TXB2 detected in the perfusing fluid were reduced in hypocapnic preparations as compared with the normocapnic ones, while the bronchoconstrictive responses to AA were not affected by CO2 tension. It is concluded that: a) biosynthesis of TXA2 is reduced in hypocapnic group if compared with that observed in normocapnic one, b) the quantitative change of AA metabolism is responsible for hypocapnia reduction of pulmonary vasoconstrictive effects of AA, c) stability of bronchoconstriction due to AA infusions in normocapnic and hypocapnic HLPs might indicate an up regulation for TXA2 bronchial smooth muscle receptors by hypocapnia.

Pulmonary arterial histology and morphometry in systemic sclerosis: a case-control autopsy study.

Morphometric measurements were performed on pulmonary arteries in 58 patients with systemic sclerosis (20 limited cutaneous and 38 diffuse cutaneous involvement [21 with and 17 without renal crisis]) and age, race, and sex matched autopsy controls. Matched pairs analysis was employed. For arteries of all sizes, the area of the intima and percent luminal occlusion were greater in the limited and diffuse (no renal crisis) groups than in controls, and these differences were statistically significant for large and medium sized vessels. The greatest luminal occlusion was found in limited cutaneous patients, and especially those with clinical evidence of pulmonary arterial hypertension, providing a rationale for the poor response to vasodilator therapy in these patients.

Prevalence of thyroid abnormalities is not increased in Alzheimer's disease.

The aim of this study was to compare the prevalence of pathologically and clinically documented thyroid disorders in patients with Alzheimer's disease (AD) and nondemented individuals. We reviewed the records of 31 patients with a pathological diagnosis of AD and 31 age-matched nondemented patients selected from autopsy files. We examined the pathological findings in the thyroid gland in both groups. Twenty-four cases (77.4%) with AD and 21 patients (67.7%) in the control group showed pathological abnormalities in the thyroid gland. The prevalence of specific types of thyroid gland abnormalities on pathological examination did not differ significantly between the AD and nondemented cases. For the 19 AD and 18 control patients in whom clinical thyroid function had been documented adequately during life, the percentage of patients with abnormal thyroid function was similar in the AD (15.7%) and control (11.1%) groups. These results do not support a relationship between AD and thyroid abnormalities.

Calcium transport and energy coupling in diabetic rat liver mitochondria.

Liver mitochondria from chronic diabetic rats took up Ca ions at a significantly slower rate than their normal counterparts. The same mitochondria, inhibited with ruthenium red, released Ca ions at a faster rate than normal mitochondria. In good agreement with the decrease of Ca ions uptake, measurement of the inner membrane potential by the safranine method yielded a significantly lower value with diabetic than with normal mitochondria. Respiratory control and P/O ratios were also decreased in diabetic mitochondria.

Hypophysectomy and adrenalectomy decrease Ca2+-induced swelling of rat liver mitochondria.

Hypophysectomized rats showed significant modifications in Ca2+-induced swelling and MgATP-induced contraction of liver mitochondria. Two-three months after operation, maximal swelling (MS), rate of swelling (VS), maximal contraction (MC) and rate of contraction (VC) were decreased by 77 (MS), 83 (VS), 79 (MC) and 62 (VC) % (in all cases P less than 0.05 as compared with normal rat mitochondria). Treatment of hypophysectomized rats with bovine pituitary extract, for two weeks, counteracted the effect of hypophysectomy, whereas treatment with bovine growth hormone was ineffective. Adrenalectomy produced similar though less extensive modification of Ca2+-induced swelling but failed to affect the MgATP-dependent mitochondrial contraction. Substrate oxidation by hypophysectomized or adrenalectomized rat mitochondria showed no modification of the energy-yielding mechanisms, capable of explaining the observed changes in mitochondrial swelling.

Renal vascular histology and morphometry in systemic sclerosis. A case-control autopsy study.

An autopsy case-control study of renal vascular histology and morphometry in systemic sclerosis (scleroderma) was performed. Thirty-five of 70 systemic sclerosis cases had renal tissue available for study: 26 had diffuse cutaneous involvement (9 with "renal crisis" and 17 without) and 9 had limited cutaneous disease (CREST syndrome [calcinosis, Raynaud's phenomenon, esophageal dysmotility, sclerodactyly, telangiectasias]). Age-matched (within 10 years) and sex-matched controls with renal specimens were obtained. New sections were cut from tissue blocks, and morphometry was completed using a Zeiss Image Analyzer. Using analysis of variance, the intimal area (Ai) was significantly increased (intimal thickening) in small and medium-sized arteries of patients with diffuse scleroderma and in small arteries of CREST patients, compared with those in controls, while a decreased medial area (Am) was seen consistently in all groups. The proportion of the vessel wall occupied by intima (Ai:[Ai + Am]) was significantly greater in all vessel size groups in patients with diffuse scleroderma compared with that in controls. The percentage of luminal occlusion was greatest in patients with diffuse disease with renal crisis. These same patients had severe edematous and mucinous intimal thickening in small and medium vessels, often in association with fibrinoid necrosis. We conclude that renal vascular structural changes are an integral part of systemic sclerosis. However, the significant differences between diffuse scleroderma patients and CREST syndrome patients, for both intimal thickening and percentage of luminal occlusion, suggest that the arterial disease in these 2 patient subsets is distinctive.

Skeletal myxoid chondrosarcoma with microtubular aggregates within rough endoplasmic reticulum.

A skeletal myxoid chondrosarcoma of the femur was examined by light and electron microscopy. Light microscopic features were characteristic for myxoid chondrosarcoma or "chordoid sarcoma." Electron microscopy disclosed crystalline arrays of microtubules within dilated rough endoplasmic reticulum, a feature previously described in extraskeletal myxoid chondrosarcomas. Tumors with the histologic characteristics of extraskeletal myxoid chondrosarcoma ("chordoid sarcoma") occurring in bone have been rarely reported. We describe herein such a tumor and document the existence of distinct microtubules within the endoplasmic reticulum.

Malignant epithelioid hemangioendothelioma of the colon. Report of a case.

Angiosarcoma of the colon with epithelioid and histiocytoid features, a malignant counterpart of epithelioid hemangioendothelioma, was observed in a 72-year-old man. The disease first manifested as a right cervical mass, with the histologic appearance of malignant, undifferentiated, large-cell epithelioid neoplasm. Light microscopy of the colonic tumor disclosed angiosarcoma, with active erythrophagocytosis and positive immunoperoxidase reactions to lysozyme, alpha-1-antitrypsin, and alpha-1-antichymotrypsin. Ultrastructural features of the tumor cells were those of intermediate between endothelial and histiocytic cells. The disease took a rapid fatal course with recurrence, peritoneal dissemination, and massive peritoneal hemorrhage. The cause remains unknown.

Cyclosporin A-induced cholestasis. The mechanism in a rat model.

Cyclosporin A (CyA) causes cholestasis in a significant proportion of transplant patients. Doses of 5, 10, and 15 mg CyA/kg body wt or the Miglyol 812 vehicle were administered intraperitoneally for 1, 2, and 3 wk to separate groups of rats to investigate the mechanism of this cholestasis. At 1 wk a dose-response relationship between serum CyA levels and increasing CyA doses was noted. A maximum CyA blood level was achieved by 2 wk with the 10- and 15-mg/kg doses. Subsequent studies were performed using the smaller (10 mg/kg) dose administered for 3 wk. This dose resulted in a marked increase in serum bile acid levels compared with vehicle-treated controls (24.6 +/- 4.0 vs. 4.3 +/- 1.2 mumol/L, p less than 0.001) without inducing significant changes in serum glutamic oxaloacetic transaminase, serum glutamic pyruvic transaminase, bilirubin, alkaline phosphatase, and albumin levels or hepatic architectural alterations. With CyA treatment, baseline bile flow decreased by 35% and bile salt secretion decreased by 25% compared with vehicle-treated animals. Cyclosporin A and vehicle-treated rats were infused intravenously with taurocholate (4 mumol/min X kg) for 2 h and then depleted of bile salts over the next 24 h. Bile samples collected over this period were graphed as bile salt secretion versus bile flow. The mean slope of the linear regression for the CyA-treated rats was 62% of the control, demonstrating a decrease in bile salt-dependent flow. Extrapolation of the linear regression to the ordinate demonstrated a 22% decrease in bile-independent flow with CyA treatment. Therefore, in our experimental model of CyA-induced cholestasis, the decrease in flow observed was the result of a decrease in both bile salt-dependent and bile salt-independent flows and occurred in the absence of significant biochemical or histologically evident hepatotoxicity.

Ultrastructural localization of rat Clara cell 10 KD secretory protein by the immunogold technique using polyclonal and monoclonal antibodies.

Using a monoclonal antibody and affinity-purified polyclonal antiserum against a 10 KD protein isolated from rat pulmonary lavage, we have localized the protein within Clara cells by a post-embedment protein A-gold technique. The gold particles were localized over the secretory granules of rat Clara cells. Ultrastructural immunolocalization was abolished when the primary antibodies were previously absorbed with purified 10 KD protein. Other pulmonary cells, including type II pneumocytes and ciliated cells, were negative with this technique. These results demonstrate the presence of the 10 KD protein in the secretory granules of the Clara cell and support the concept that this protein constitutes a specific and unique secretory product of Clara cells.

Effect of diabetes, adrenalectomy and hypophysectomy on D-3-hydroxybutyrate dehydrogenase activity and substrate oxidation by rat mitochondria.

In good agreement with previous observations, D-3-hydroxybutyrate dehydrogenase (OHBD) activity in disrupted liver mitochondria decreased significantly in chronic diabetic rats, whereas insulin treatment prevented this decrease. Similar effects were observed with kidney mitochondria. Adrenalectomy increased OHBD activity in liver and brain mitochondria from normal rats and in liver mitochondria from diabetic rats. Corticosterone depressed OHBD activity in liver mitochondria of normal rats and restored the normal level of OHBD in adrenalectomized rats. Hypophysectomy increased OHBD activity in normal rats, this effect being further increased by growth hormone administration. D-3-hydroxybutyrate (OHB) oxidation by liver mitochondria from diabetic, adrenalectomized or hypophysectomized rats varied as expected from OHBD activity in disrupted mitochondria, but to a relatively lesser degree. Submitochondrial particles from diabetic rats showed significant diminution in OHB-, succinate- and NADH-oxidase activities, whereas mitochondria from diabetic rats oxidized succinate and L-malate + L-glutamate at a slower rate than the corresponding controls.

Arachidonic acid and pulmonary function in heart-lung-preparation of guinea-pig: modulation by PCO2.

In normocapnic and deeply hypocapnic guinea-pig heart-lung-preparations (HLPs), dose-response relationships were estimated for the bronchoconstrictor and pulmonary hypertensive responses to histamine (H), 5 hydroxytryptamine (5HT), arachidonic acid (AA) and U-46619, a prostaglandin endoperoxide analogue acting on thromboxane (TXA2) receptors. Hypocapnia potentiated in a different way the bronchoconstrictor effects of AA (increased slope of dose-response curve) and of U-46619 (shift to the left of the curve). The pulmonary vascular effects of U-46619 were unaffected by CO2 tension, whereas a linear log dose-dependence of the pulmonary hypertensive responses to AA was present only in hypocapnic HLPs. The amount of TXA2-like material released in normocapnic HLPs was compatible with the AA/U-46619 potency ratio calculated for the bronchoconstrictor responses in normocapnic HLPs and for the pulmonary vascular responses in hypocapnic HLPs. The above described effects of hypocapnia were different from those produced on the bronchial and pulmonary vascular reactivity to H and 5HT, suggesting that specific mechanisms are involved in the modulating effect of PCO2. The inhibition by indomethacin of AA-induced pulmonary vasoconstriction was unaffected by changes in CO2 tension; conversely, the bronchoconstrictor effects of AA were more substantially reduced by indomethacin in normocapnic HLPs. It is concluded that: the relative contribution of different AA metabolites to the final response of the airway system to the precursor is affected by changes in CO2 tension; different receptorial or prereceptorial mechanisms are involved in the CO2-AA interaction taking place in the two components of the lung parenchyma; the pulmonary outflow of AA metabolites provides only circumstantial evidence of the functional meaning of this release.

Primary adenocarcinoma arising at an ileostomy site. An unusual complication after colectomy for ulcerative colitis.

A case is reported of a 45-year-old man who developed a primary adenocarcinoma at the ileostomy site 23 years after proctocolectomy for ulcerative colitis. The patient underwent wide local excision of the tumor and died 13 months later with disseminated adenocarcinoma. Including the present case, a total of seven patients with ileostomy adenocarcinoma have been reported in the literature. Six patients were treated surgically for ulcerative colitis, and the other one for adenomatous polyposis coli. The diagnosis of stomal malignancy was made three to 23 years after the ileostomy (mean interval, 13 years). Of the four patients available for clinical follow-up, two died of disseminated adenocarcinoma ten and 13 months after diagnosis. Awareness of this unusual but important complication of ileostomy should lead to earlier diagnosis and treatment with improvement in prognosis.