Role of synaptophysin in the intraoperative assessment of quadrantic innervation of the proximal doughnut in Hirschsprung disease.

BACKGROUND: Symptoms may persist in a retained aganglionic segment of the colon after corrective (pull-through) surgery in Hirschsprung disease (HD). Thus, it is important to assess the proximal doughnut for innervation abnormalities intraoperatively by frozen sections stained with conventional haematoxylin and eosin stain and supported by rapid acetylcholinesterase (AChE) histochemistry. When the doughnut is proximal to the sigmoid colon, AChE is not useful and requires ratification by yet another rapid technique and hence this study. METHODS: Two pathologists independently evaluated fresh doughnuts from the proximal bowel clinically assumed to be of normal innervation intraoperatively and chosen for anastomosis in patients with HD along with controls using AChE and synaptophysin (SY) immunohistochemistry. RESULTS: From 38 patients with HD, 28 doughnuts (63.7%) showed normal innervation with intense SY activity in the mucosa, the muscularis and the ganglion cells. The circumferential aganglionic doughnuts (abnormal innervation) (n= 6, 13.6%) showed neither SY-positive fibres in the mucosa nor in the muscularis. The abnormal transition zone doughnuts (n=10, 22.7%) showed involvement of three quadrants of the doughnut in one, two quadrants in three and one quadrant in six with decreased SY-positive fibres in the muscularis and scattered ganglion cells with a statistically significant measure of agreement of (κ=0.973) between the two. CONCLUSION: The pattern, intensity and distribution of SY-positive fibres in the muscularis propria of the doughnut of the proximal bowel chosen intraoperatively for anastomosis in HD can identify sectors with abnormal innervation allowing the surgeon to seek normal innervation status more proximally to avoid complications.

The distal level of normally innervated bowel in long segment colonic Hirschsprung's disease.

PURPOSE: The definitive surgical management of Hirschsprung's disease (HD) depends on an unambiguous histological diagnosis of bowel aganglionosis, as well as an accurate assessment of its proximal extent from the rectum or 'leveling'. This study aimed to evaluate and compare the transition zone in rectosigmoid and long segment colonic forms of HD. METHODS: One hundred and fifteen cases of rectosigmoid Hirschsprung's disease (RSHD) and 20 cases of long segment colonic Hirschsprung's disease (LScHD) managed from 1998 to 2008 with a consensus prospective protocol were included in this clinicopathological analysis. RESULTS: In RSHD, the radiologic transition zone (rTZ)/gross appearance at laparotomy correlated with the histological 'leveling' in the majority (96%); the histologic transition zone was also short and predictable. In contrast, the rTZ in LScHD was conclusive in only 53%; overall the radiologic and histological transition zones concurred in only 26%. The histologic transition zone extended proximally for an unpredictable length before normal circumferential innervation was noted. CONCLUSIONS: A stringent intraoperative histological evaluation of the colon as outlined is more helpful than radiology to establish the distal level of normally innervated bowel in LScHD and achieve an accurate leveling during a colostomy/pull through.

Ocular spherulocystosis.

Spherulocystosis (myospherulosis), a rare entity, is characterised by histopathological changes of sac-like refractile structures in soft tissue and presents clinically as an inflammatory lesion. It is caused by trauma or is induced iatrogenically by the topical application of fat based ointments. A case of myospherulosis confined to the region of the lacrimal caruncle, which developed soon after topical antibiotic application for suspected conjunctivitis, in a 49 year old woman is reported. Though this lesion is uncommon, this report illustrates the importance of recognising this condition to avoid confusion with and treatment for fungal infection.

Simultaneous separation and quantitation of four antiepileptic drugs--a study with potential for use in patient drug level monitoring.

The purpose of this study was to illustrate the applicability of high-performance liquid chromatography (HPLC) coupled with evaporative light scattering detector (ELSD) in simultaneously separating and quantitating four commonly used antiepileptic drugs (AEDs). A mixture of the four AEDs were separated using a C8 column using volatile mobile phases and were detected using ELSD. Optimal instrumental conditions were obtained by assessing the effect of various critical experimental parameters such as evaporator tube temperature, carrier gas flow rate, photomultiplier gain on separation efficiency, accuracy, reproducibility and sensitivity of measurement on all four AEDs. A novel, rapid, accurate, sensitive, reproducible and robust HPLC-ELSD method for simultaneous separation and quantitation of four commonly used AEDs was developed. The physical basis of the results obtained as a consequence of varying several critical experimental parameters has been explained. This study illustrates the potential for use of HPLC-ELSD in drug level monitoring of patients undergoing mono- or polytherapy for epilepsy.

A modified technique for the diagnosis of Hirschsprung disease from rectal biopsies.

BACKGROUND: A diagnosis of Hirschsprung disease requires the demonstration of acetylcholinesterase fibres on frozen sections obtained from snap frozen biopsies of the rectum. This histochemical technique is generally not available in laboratories in developing countries. We improvised on the methodology of tissue preservation to make the staining technique more user-friendly, economical and reliable in demonstrating acetylcholinesterase activity in fresh rectal mucosal biopsies for the diagnosis of Hirschsprung disease. METHODS: Between June 1999 and May 2002 fresh rectal biopsies from 40 suspected cases of Hirschsprung disease were processed for routine frozen section (not snap frozen by liquid nitrogen) and stained by the Karnovsky and Roots method. These sections were assessed for the staining pattern of acetylcholinesterase fibres. The thickness of the nerve fibres and muscularis mucosa was assessed morphometrically. These were compared with biopsies obtained from 6 age-matched controls undergoing surgery for unrelated complaints. RESULTS: The sections stained for acetylcholinesterase by this improvised method of tissue fixation were good and crisp. A definite diagnosis of Hirschsprung disease was made in 25 cases and intestinal neuronal dysplasia in 1. The remaining 14 cases showed an equivocal staining pattern with no hypertrophic nerve bundles, thus excluding a diagnosis of Hirschsprung disease. The mean thickness of the submucosal nerve trunks measured in these enzyme-stained sections was found to be inversely proportional to the mean thickness of the muscularis mucosa. CONCLUSION: Our study on cryostat-cut sections suggests an inverse relationship between the thickness of the muscularis mucosa and the calibre of the nerve trunk--thinner the nerve trunk, thicker the muscularis mucosa and vice versa. Also, routine frozen sections, instead of snap frozen ones taken from a fresh rectal biopsy and stained by the Karnovsky and Roots method for acetylcholinesterase activity, are reliable for the diagnosis of Hirschsprung disease and are within the capability of a simple histopathology laboratory in a developing country.

Combined hepatocellular and cystadenocarcinoma presenting as a giant cyst of the liver--a case report.

Primary cystic lesions of the liver are very rare. Most of the solid tumours are hepatocellular carcinomas (HCC) with a smaller number being cholangiocarcinomas. The association of HCC with other primary liver malignancies is also extremely rare. This case report is about a 27 year old male patient who presented with a giant cystic lesion of the left liver. A CT scan showed a cystic lesion with internal septations and a thrombus in the main portal vein. The patient underwent an extended left hepatectomy and a portal venotomy with removal of the thrombus. Coexistent hepatocellular and cystadenocarcinoma were reported on histopathological examination. The patient was put on 5-FU postoperatively. He is doing well 11 months after surgery.

Choriocarcinoma of the ovary in a post-menopausal woman.

A case of a Choriocarcinoma of the Ovary has been reported in a 50 year old post-menopausal woman. At laparotomy, the lesion was seen as a large necrotic ovarian mass. The problems of distinguishing Gestational from Non-gestational ovarian choriocarcinoma are discussed. Cytogenetic studies are indicated to investigate potential reasons for the difference in prognosis.

Isolated necrotising arteritis of the cervix and myometrium: a much neglected but puzzling entity.

Arteritis of the uterine cervix and corpus described here was an incidental finding at the routine histopathological examination of the hysterectomy specimen resected from a 62 year old female who underwent laparotomy for twisted ovarian cyst. Investigations and eleven months of follow up without any specific treatment for arteritis, have shown no systemic involvement. This case highlights that a knowledge of such isolated arteritis is of importance to the physician to avoid misdiagnosing it as polyarteritis nodosa and treat with systemic steroids.

Malignant pheochromocytoma with cutaneous metastases presenting with hemolytic anemia and pyrexia of unknown origin.

We describe a 43 year old female who presented with pyrexia of unknown origin associated with Coomb's negative hemolytic anemia and impaired liver function tests of six months duration. A routine abdominal computerised tomographic scan showed a mass in the left adrenal which was excised at laparotomy and histologically diagnosed as pheochromocytoma. The hemolysis continued to worsen with development of resistance to steroid therapy. Subsequently she developed multiple firm to hard painless cutaneous nodules which were aspirated for cytologic examination. The cytology picture was that of a neuroendocrine tumour with cell morphology similar to that of the adrenal pheochromocytoma. Such an association of hemolytic anemia, pyrexia and cutaneous metastases in a case of malignant pheochromocytoma has not been described earlier in the medical literature.

Megacystis microcolon intestinal hypoperistalsis syndrome.

Megacystis microcolon intestinal hypoperistalsis syndrome (MMIHS) is a rare cause of intestinal obstruction mainly affecting female neonates. We present a case of a newborn female infant with a history of abdominal distension, bilious vomiting and decreased urine output. Barium enema showed a microcolon. Patient died soon after admission and the autopsy revealed a shortened bowel, a microcolon with abundant ganglion cells in the myenteric plexus, and an enlarged urinary bladder. An interesting finding in this case was the presence of enlarged nerve bundles containing several large ganglion cells on the lateral wall of the cervix. The salient clinical and autopsy findings in this case are presented.

Intestinal neuronal dysplasia type A.

Intestinal neuronal dysplasia is a rare cause of bowel obstruction in neonates and infants, the diagnosis of which poses a formidable challenge to both clinicians and pathologists alike. The importance of this entity lies not only in the fact that it mimics Hirschsprung's disease, but also in that untreated cases, particularly of type A, may prove fatal. The authors describe one such case of intestinal neuronal dysplasia of type A, which was diagnosed at autopsy.

Hirschsprung's disease, associated rare congenital anomalies.

OBJECTIVE: Hirschsprung's disease may be associated with a number of congenital anomalies of which Down's syndrome and intestinal atresias are commonly encountered. The study aimed to assess the impact of rare associated anomalies on the diagnosis and management of Hirschsprung's disease. METHODS: A retrospective review of the clinical presentation, diagnosis and outcome of thirty five consecutive newly diagnosed cases of Hirschsprung's disease encountered over two years was performed. RESULTS: Besides Down's syndrome (two), intestinal atresia (one) and pigmentary ocular defects (two), three rare anomalies (Occipital meningocele, Calcific meconium cyst with anal stenosis, Malrotation) were encountered in four of thirty five cases. The clinical features, radiologic anatomy and gross morphology of the bowel were unconventional and the diagnosis was supported by intraoperative acetylcholinesterase staining of biopsies. Though the diagnosis was relatively delayed in these cases, the outcome has been comparable to the rest. The dilemma in their diagnosis and management and their possible pathoembryology is discussed. CONCLUSION: Awareness of such associations and a specific investigative protocol is imperative for timely diagnosis and minimal morbidity in complex presentations of Hirschsprung's disease.

A rare cause of massive lymphadenopathy.

Rosai Dorfman Disease is a rare disease presenting mainly as painless enlargement of cervical lymph nodes. It is a benign disorder showing unique histological features characterized by reactive proliferation of sinus histiocytes. In 40% of cases, extranodal sites may be involved. The condition has to be differentiated from certain types of B cell lymphoma showing similar features. We report here a 17 year old male presenting with nodal and extranodal involvement of Rosai Dorfman Disease.

Plasmacytoma of larynx--a case report.

Plasma cell myeloma, the most common plasma cell neoplasm, is characterized by the presence of multiple lesions in the bone marrow. A single isolated lesion may occur either in bone (solitary plasmacytoma of bone) or in soft tissue (extramedullary plasmacytoma). Most cases of extramedullary plasmacytoma occur in the head and neck region. The diagnosis is established by histopathology and immunohistochemistry. A detailed evaluation for lesions at other sites is recommended as extramedullary plasmacytoma treated by radiation therapy has better survival rates than plasma cell myeloma, which is treated by chemotherapy. A case of plasmacytoma of the larynx is presented highlighting clinical and histological features with a review of literature.

Thymic carcinoma with glandular differentiation arising in a congenital thymic cyst.

A case of thymic carcinoma with glandular differentiation arising in congenital thymic cyst in a 50-year-old male is reported. Neoplasia in congenital thymic cyst is a rare phenomenon and is seldom associated with glandular differentiation. It is important to differentiate these neoplasms from carcinomas of the lower respiratory tract and from thymic carcinomas with secondary cystic degeneration, in view of their relatively good prognosis and the therapeutic implications.

Toward the development of an injectable dosage form of propofol: preparation and evaluation of propofol-sulfobutyl ether 7-beta-cyclodextrin complex.

The objectives of the present study were to undertake activities toward the development of an aqueous-based formulation of propofol (2,6-diisopropyl phenol), using sulfobutylether 7-beta-cyclodextrin (SBECD). Preformulation studies, including high performance liquid chromatography (HPLC) method development and phase-solubility evaluation in the presence of SBECD were conducted. It was determined that equilibrium solubility has been reached by 4-day and 7-day phase-solubility analysis at 30 degrees C and 37 degrees C. The apparent binding constants and various thermodynamic parameters were calculated from this data. These results suggest that "nonclassical hydrophobic effects" are the driving forces for inclusion complex formation. Compounding and lyophilization of the formulation with 20% SBECD yielded a product with propofol concentration of 10 mg/mL. The formulation properties were probed by using techniques that included modulated differential scanning calorimetry (MDSC) and Karl Fischer analysis. MDSC showed that propofol, SBECD, and the Propofol-SBECD complex displayed thermal properties at widely varying temperatures, suggesting the formation of a new solid form. The active pharmaceutical ingredient in the liquid formulation and lyophilized product was determined by the newly developed and qualified HPLC method. Short-term stability studies of the liquid formulation showed that they were stable for a month at 4 degrees C. Short-term stability studies of the freeze-dried cakes showed that the product was stable for over a month at 4 degrees C, 37 degrees C, and 50 degrees C. Based on these preliminary results, we believe that an aqueous based injectable formulation of propofol with sulfobutylether 7-beta-cyclodextrin can be successfully developed.

Generation and fate of enamines in the microsomal metabolism of cyclic tertiary amines.

Microsomal oxidation of 1-benzylpiperidine (1-BP) and its cis-2,6-dimethyl analog was studied to assess the involvement of endocyclic enamines, in equilibrium with the initially formed iminiums, in the metabolic activation of cyclic tertiary amines such as phencyclidine. Since the iminiums can be trapped with cyanide, the selective prevention by cyanide of the metabolic production of 1-benzyl-3-piperidone from 1-BP implicates the iminium in equilibrium with enamine as the source of this metabolite. In cases where iminium-enamine coupling is sterically prevented, the iminium in equilibrium with enamine species can be studied independently and are found to be more potent metabolism-dependent inactivators of cytochrome P-450 than are the corresponding parent amines. Possible mechanisms for biological oxidation of cyclic enamines to reactive intermediates are considered.

Haemoprotein-mediated metabolism of enamines and the possible involvement of one-electron oxidations.

1. Microsomal metabolism of 1-benzylpiperidine (1-BP), its cis-2,6-dimethyl (cis-2,6-DMBP), 4,4-dimethyl (4,4-DMBP), and alpha, alpha-dimethyl (alpha, alpha-DMBP) analogues, and phencyclidine (PCP) has been studied to assess the involvement of P450 oxidation of the enamine tautomers of the initial endocyclic iminium metabolites. 2. The selective prevention by cyanide of the metabolite production of 1-benzyl-3-piperidone but not 1-benzyl-3-piperidinol from 1-BP is consistent with the enamine as the source of the 3-one metabolite. 3. The parent amines and particularly the independently prepared iminium species induced a pattern of metabolism-dependent irreversible inactivation of P450 benz-phetamine demethylase activity, consistent with involvement of enamine C-3 oxidation in the inactivation process. 4. Substrate activity of the endocyclic enamines and alpha-aminoketones (presumably the enol-enamine tautomers) for horseradish peroxidase under conditions where simple aliphatic amines display no activity is consistent with metabolic one-electron oxidations of the enamines.