Association of gastric fluid microbes at birth with severe bronchopulmonary dysplasia.

OBJECTIVE: Gastric fluid microbes were examined in preterm infants at birth to assess their influence on the postnatal outcome. STUDY DESIGN: Prospective cohort study. SETTING: Level III neonatal intensive care unit. PATIENTS: A total of 103 premature neonates with a gestational age of less than 32 weeks. MAIN OUTCOME MEASURE: Gastric fluid microbes were identified by analysis of bacterial 16S ribosomal RNA gene. Additionally, the urease gene of Ureaplasma species was detected by polymerase chain reaction of gastric fluid obtained at birth and/or tracheal aspirate from ventilated preterm infants. The association between detection of microbes and bronchopulmonary dysplasia was investigated through assessment from clinical features and by a lung injury marker (KL-6). RESULTS: Forty-two of 103 gastric fluid specimens were positive for microbes. Ureaplasma species were detected in 23 of the 42 (55%) gastric fluid specimens. All infants with Ureaplasma species in tracheal aspirate fluid also had positive gastric fluid specimens. Compared to infants negative for gastric fluid microbes, infants positive for microbes had higher rates of maternal chorioamnionitis (18% vs 78%), premature rupture of membranes (11% vs 55%), severe bronchopulmonary dysplasia (1.6% vs 14%) and showed higher plasma KL-6 levels during the initial 4 weeks of life. CONCLUSION: Detection of gastric fluid microbes was correlated well with antenatal infection and severe bronchopulmonary dysplasia. Detection of Ureaplasma species in gastric fluid was associated with subsequent respiratory colonisation. These results suggest that antenatal exposure of the immature fetus to microbes may cause lung injury and promote the onset of bronchopulmonary dysplasia.

Alpha-1-antitrypsin deficiency (Siiyama) as indication for lung transplantation: proper timing for surgical intervention.

The authors report a new familial case of alpha-1- antitrypsin (AAT) deficiency with severe pulmonary emphysema and hemoptysis. A severely reduced serum AAT level of the proband, a 56-year-old farmer's wife and her sister were observed. Mutation analysis of the AAT gene was performed using allele-specific polymerase chain reaction (PCR) analysis followed by direct sequencing. The proband and her younger sister proved to be homozygous for PISiiyama. Although home oxygen therapy was induced in addition to previous medications including bronchodilators and cardiovascular conditioning, the proband's rate of decline of forced expiratory volume at one second (FEV1) was progressing. Lung transplantation was therefore advisable for the patient. Clinical analysis on Japanese cases reported in the literature shows that the rate of decline of FEV1 is one of the most convenient prognostic factors to find proper timing for surgical intervention. Lung transplantation is one of the best reliable current therapies to improve quality of life of severely impaired patients.

Presence of perivenular elastic fibers in nonalcoholic steatohepatitis Fibrosis Stage III.

Elastic fibers appear in extensive old fibrotic foci in general. We examined an association between hepatic fibrosis stage and the presence of perivenular elastic fibers in nonalcoholic steatohepatitis (NASH). A total of 48 liver needle biopsy specimens were used, taken from 48 cases with NASH. Fibrosis Stage (Brunt E, et al. Am. J. Gastroenterol. 1999) of the cases was as follows; six Fibrosis Stage I, twenty-two Fibrosis Stage II, and twenty Fibrosis Stage III. We examined Orcein stain sections in all of the liver needle biopsy specimens. In all twenty Fibrosis Stage III cases, perivenular elastic fiber bundles were observed. In contrast, perivenular elastic fibers were detected only in one of the six Fibrosis Stage I and two of the twenty-two Fibrosis Stage II cases. In liver needle biopsy specimens of NASH, detection of perivenular elastic fibers is useful in deciding Fibrosis Stage III.

Renal leiomyoma: an immunohistochemical, ultrastructural and comparative genomic hybridization study.

Renal leiomyoma is a rare neoplasm. We report such a case in a 57-year-old Japanese woman who was found to have a mass in the left kidney. The histological examination disclosed the proliferation of spindle cells showing a benign appearance. Entrapped tubular cells were observed in the peripheral area of the tumor. The immunohistochemical examination of spindle neoplastic cells showed a positive reaction for alpha smooth muscle actin, h-caldesmon, l-caldesmon, calponin, muscle actin, myosin and desmin. Additionally, the ultrastructural examination of the tumor showed membrane caveolae and myofilaments in the cytoplasm. This tumor was considered to show a differentiation into smooth muscle cells. The comparative genomic hybridization of the tumor detected the combined losses of chromosomes 4, 6, 12 and 14 which has not been previously described in renal tumors. Finally, the immunohistochemical panel of smooth muscle markers and ultrastructural and genetic study may be useful in diagnosing renal leiomyoma.

The distribution of myofibroblasts and CD34-positive stromal cells in normal renal pelvis and ureter and their cancers.

In this article, we examined the distribution of myofibroblasts and CD34-positive stromal cells in normal renal pelvis and ureter and their cancers using immunohistochemistry. Eighteen tumors and normal tissues apart from the main tumor were examined. In the wall of normal renal pelvis and ureter, no myofibroblasts were observed through all layers, but CD34-positive stromal cells were observed in the deep area of lamina propria, muscular layer and adventitia. In the stroma of renal pelvic and ureteral cancers, myofibroblasts were distributed in fifteen tumors and were absent in three tumors. All three tumors containing no myofibroblasts in the stroma were non-invasive type and all invasive cancers contained myofibroblasts in the stroma. CD34-positive stromal cells were consistently absent in the stroma of cancers, irrespective of the invasiveness. Finally, myofibroblasts are major stromal components in renal pelvic and ureteral cancers, particularly in invasive cancers, and CD34-positive stromal cells are consistently absent or lost in the stroma of their cancers. These findings suggest that the invasion of renal pelvic and ureteral cancers may cause the phenotypic change of stromal cells.

Retroperitoneal hemorrhage due to bilateral adrenal metastases from lung adenocarcinoma.

A 56-yr-old man was admitted to our university hospital for severe back pain one month after a resection for lung adenocarcinoma (stage IIIA) without evidence of the adrenal mass. Computed tomography (CT) of the abdomen showed bilateral bleeding of adrenal tumors. Endocrinological laboratory studies showed high plasma ACTH and normal serum cortisol levels with the loss of circadian rhythm. Although plasma ACTH levels increased, there was no cortisol response to administration of human corticotropichormone (hCRH). Core-needle biopsy was performed on the right adrenal tumor and revealed adenocarcinoma cells mimicking a primary lung tumor previously examined. We diagnosed retroperitoneal hemorrhage due to bilateral adrenal gland metastasis from lung adenocarcinoma with adrenal insufficiency. Adrenal metastases most commonly originate from a primary lung tumor, followed by stomach, esophagus and liver/bile ducts. Bilateral adrenal metastases were noted in approximately half of all adrenal metastases patients. Clinically significant adrenal hemorrhage by metastasis is exceedingly rare and non-specific symptoms, such as abdominal, chest or back pain, nausea and vomiting, confusion, weakness, hypotension, shock and high fever, are often observed in these patients. We present a case of massive retroperitoneal hemorrhage and adrenal insufficiency due to adrenal gland metastasis from adenocarcinoma of lung.

Cytokeratin-positive subserosal myofibroblasts in gastroduodenal ulcer; another type of myofibroblasts.

To investigate the distribution and origin of alpha-smooth muscle actin (ASMA)-positive stromal cells in the perforation of human gastroduodenal ulcers. Perforative lesions of 24 surgically resected gastroduodenal ulcers were examined immunohistochemically for ASMA, HCD, CD34, CD31, CAM5.2 and HMW-CK, and double staining of ASMA and CAM5.2 was also performed. In addition, to determine the cell source of collagen, in situ hybridization of collagen I mRNA was performed. In the normal gastroduodenal wall, the reticular network of CD34-positive stromal cells was identified in the muscularis mucosa, submucosa, muscular propria, and subserosa. In the subepithelial area, many myofibroblasts were observed, whereas no CD34-positive stromal cells were seen. In areas neighboring ulcerative lesions, no CD34-positive stromal cells were observed, but a significant number of myofibroblasts were present there. In the deep layer of ulceration, numerous fusiform or stellate stromal cells strongly positive for ASMA and CAM5.2 were observed in the subserosal area around the perforation. In the same site, many cells co-expressing ASMA and CAM5.2 were identified by double staining. In contrast, in the surface layer of ulceration, stromal cells expressing only ASMA were observed. The cytokeratin-positive subserosal myofibroblastic cell in human gastroduodenal ulcer is a novel type of myofibroblast.

The distribution pattern of myofibroblasts in the stroma of human bladder carcinoma depends on their invasiveness.

The presence of myofibroblasts has been elucidated in the stroma of neoplasm of various organs. In the present article, we studied the distribution of myofibroblasts in the stroma of bladder carcinoma. Twenty-five surgical resected bladder tumors (urothelial carcinoma, n = 21; combined urothelial carcinoma and adenocarcinoma, n = 2; sarcomatoid squamous cell carcinoma, n = 1; combined urothelial carcinoma and squamous cell carcinoma, n = 1) were selected and we evaluated the distribution of myofibroblasts using immunohistochemical, electron and immunoelectron microscopic techniques. Immunohistochemically, the distribution pattern of myofibroblasts in invasive and non-invasive carcinomas were predominantly fascicular and reticular forms, respectively. Moreover, myofibroblasts around bladder carcinoma cells were confirmed by electron microscope. Understanding the distribution pattern of myofibroblasts in the stroma of bladder carcinoma may provide available information about the presence of carcinoma invasion.

Intranasal administration of ACTH(1-24) stimulates catecholamine secretion.

Previously, we reported that intranasal (IN) ACTH(1-24) administration stimulates adrenocortical steroid secretion in normal subjects. To determine the efficiency of transmucosal absorption of ACTH into the adrenal medulla, we measured serum cortisol, aldosterone, epinephrine, norepinephrine and dopamine levels after IN vs. intravenous (IV) administration of 250 microg ACTH(1-24) in 7 healthy adult men (mean age 21.7 +/- 1.2 yr; range, 21 - 24 yr). Blood was collected at 0, 30, 60 and 120 min after administration of ACTH(1-24), and the levels of adrenocortical steroids and catecholamines were measured by specific RIA and HPLC methods, respectively. There were no side effects associated with IN or IV ACTH administration. Consistent with the previous study, serum cortisol and aldosterone increased after IN administration of ACTH(1-24), peaking 30 min after administration. Sixty minutes after IN and IV administration of ACTH, epinephrine levels increased by 41.9 +/- 13.1 % and 63.3 +/- 11.8 %, respectively, and remained elevated throughout the sampling period. Thirty minutes after IN or IV administration of ACTH(1-24), plasma norepinephrine levels increased by 55.9 +/- 13.4 % and 73.7 +/- 15.0 %, respectively, peaking 30 min after ACTH(1-24) administration, and decreasing to basal levels within 60 min. Plasma dopamine levels did not change after IN administration of ACTH(1-24). Adrenocortical steroid and catecholamine levels did not increase after IN administration of saline. These results demonstrate that IN administration of ACTH(1-24) not only stimulates adrenocortical steroids, but also epinephrine and norepinephrine.

Pulmonary adenocarcinoma with micropapillary component: an immunohistochemical study. Case report.

Micropapillary carcinoma has been described in various organs, including the breast, urinary bladder, ovary and lung. We here present a case of pulmonary micropapillary carcinoma in a 72-year-old Japanese man who died of respiratory failure and septic shock, following which autopsy was performed. A mass measuring 2.5 x 2.5 x 2.5 cm was observed in the left lower lobe of the lung. The tumor showed moderately differentiated papillary adenocarcinoma with a focal micropapillary component. Carcinomatous lymphangiosis was also observed in the left lung and metastatic lesions were observed in the bilateral lung, liver, vertebra, muscle layer of the urinary bladder, right adrenal gland, spleen and lymph nodes. The micropapillary component was predominant at some metastatic sites. Immunohistochemically, both the adenocarcinoma and micropapillary components were positive for cytokeratin (CK) 7, CK19, TTF (thyroid transcription factor)-1, carcinoembryonic antigen (CEA) and surfactant apoprotein A (SP-A), and negative for CK20, estrogen receptor, progesterone receptor, uroplakin III, and CA125. The invasive area of the conventional adenocarcinoma component contained a large number of myofibroblasts, whereas the stroma of the micropapillary component contained a small number of myofibroblasts. However, no myofibroblasts were observed in the stroma of the central core of the non-invasive micropapillary carcinoma. Several lymphatic invasions by neoplastic cells were identified in the peripheral area of the micropapillary component using D2-40 antibody. The immunohistochemical profile may be helpful in determining the primary location of the neoplasm containing micropapillary features. Myofibroblasts are present in the stroma of the invasive neoplastic nests in the micropapillary component as well as the conventional adenocarcinoma component, and D2-40 monoclonal antibody may be useful for evaluating the lymphatic invasion of pulmonary micropapillary carcinoma.

Consistent lack of CD34-positive stromal cells in the stroma of malignant breast lesions.

To examine the distribution of CD34-positive and ASMA-positive stromal cells in various breast lesions, we performed immunohistochemical assays (using a streptavidin-biotin immunoperoxidase technique) of tissue specimens, obtained by excisional biopsy and partial or total mastectomy, from 62 patients with breast lesions. Specimens were obtained from 64 lesions as follows: fibrocystic disease (n=12), intraductal papilloma (n=4), fibroadenoma (n=17), invasive lobular carcinoma (n=6), invasive ductal carcinoma (n=20) and invasive micropapillary carcinoma (n=5). In normal breast tissue (controls), CD34-positive spindle cells were abundant in the intralobular stroma, but no ASMA-positive stromal cells were identified except myoepithelial cells. Small to large numbers of CD34-positive cells were observed in the stroma of 29 of 33 benign diseases. In all invasive carcinomas (lobular, ductal and micropapillary), no CD34-positive stromal cells were observed in the stroma. In the stroma of benign lesions, the number of ASMA-positive stromal cells was various, but the stroma of all invasive breast cancers contained ASMA-positive stromal cells. The present results indicate that disappearance of CD34-positive stromal cells consistently occurs in the stroma of invasive carcinoma of the breast, irrespective of histological type and may be associated with the presence of ASMA-positive stromal cells.

The distribution of CD34-positive stromal cells and myofibroblasts in colorectal carcinoid tumors.

In order to understand the stromal reaction associated with colorectal neoplasms, we examined specimens from 26 patients including normal colorectal tissues (n=15), carcinoid tumors (n=12), well differentiated adenocarcinomas (n=10), and poorly differentiated adenocarcinomas (n=4), using an immunohistochemical method. Myofibroblasts and CD34-positive stromal cells were distributed in the mucosa and in the area between the submucosal and subserosal layers, respectively. However, the distribution of these cells markedly changed with the invasion of neoplasms. Namely, myofibroblasts were abundant in the invasive stroma of all colorectal neoplasms. CD34-positive stromal cells were completely absent from the invasive stroma of colorectal cancers. On the other hand, CD34-positive stromal cells were absent from four out of five carcinoid tumor cases with lesions measuring less than 2 mm in size, but were present in all seven cases of carcinoid tumors measuring more than 2 mm. Double-immunostaining identified stromal cells expressing both ASMA and CD34 in several carcinoid tumor cases. Finally, no CD34-positive stromal cells were observed in the invasive stroma of colorectal cancers. However, the distribution of these cells in carcinoid tumors may depend on the lesion size. Namely, CD34-positive stromal cells existed between neoplastic nests in large-sized carcinoid tumors. Myofibroblasts in the stroma of colorectal neoplasms may originate from CD34-positive stromal cells.

The participation of myofibroblasts in the capsular formation of human conventional and chromophobe renal cell carcinomas.

The presence of myofibroblasts has been elucidated in neoplastic capsules of various organs. In the present article, we examine the presence of myofibroblasts in the capsule of renal cell carcinoma (RCC) and discuss the origin of the myofibroblasts. Nineteen renal tumors (conventional RCC, n=17; chromophobe RCC, n=2) with evident and totally surrounded fibrous capsule were selected. Abundant myofibroblasts were immunohistochemically observed in the capsule of the RCCs. These findings were confirmed by electron and immunoelectron microscopic studies of three conventional RCCs. Type III and I collagens were predominant in the outer and inner layers of the RCC capsule, respectively. The cytoplasm of the tubular epithelial cells in the tissue surrounding the neoplastic capsule stained positively for transforming growth factor (TGF)-beta 1. In situ hybridization detected type I collagen mRNA in myofibroblasts of the capsule. Myofibroblasts may participate in the capsular formation of conventional and chromophobe RCCs through the collagen production.

The appearance of myofibroblasts and the disappearance of CD34-positive stromal cells in the area adjacent to xanthogranulomatous foci of chronic cholecystitis.

We investigated the distribution of myofibroblasts and CD34-positive stromal cells in normal gallbladder and its pathological conditions (cholecystitis, n=25) using immunohistochemistry and in situ hybridization. In the wall of normal gallbladder, myofibroblasts were generally absent from all layers, but many CD34-positive stromal cells were observed in the connective tissue layer. In chronic cholecystitis with mild perimuscular fibrosis, a small to moderate number of myofibroblasts appeared in the mucosal layer. In chronic cholecystitis with marked perimuscular fibrosis, a small to large number of myofibroblasts appeared predominantly in the connective tissue layer, whereas the number of CD34-positive stromal cells decreased at the same location, although the number of myofibroblasts increased. In chronic cholecystitis with xanthogranulomatous foci, a small to large number of myofibroblasts were observed in the periphery of the xanthogranulomatous reaction and adjacent area. In contrast, CD34-positive stromal cells were completely absent or were limited to the area just around the xanthogranulomatous reaction. Induction of collagen type I and III mRNA was predominantly observed in the cytoplasm of myofibroblasts associated with the marked fibrosis, which consisted primarily of mature collagen fibers, and in the cytoplasm of myofibroblasts around the xanthogranulomatous reaction, respectively. Finally, myofibroblasts were observed in all subtypes. The increased number of myofibroblasts was most prominent in the connective tissue layer of chronic cholecystitis with marked perimuscular fibrosis or in the area adjacent to xanthogranulomatous foci of chronic cholecystitis. Under these conditions, CD34-positive stromal cells tended to disappear from the connective tissue layer, which exhibited an increase in myofibroblasts.

Review of mucinous tubular and spindle-cell carcinoma of the kidney with a focus on clinical and pathobiological aspects.

Recently, the characterization of mucinous tubular and spindle-cell carcinoma (MTSCC) has been established. MTSCC predominantly occurs in females. This tumor is histologically characterized by eosinophilic cytoplasm, elongated and anastomosing tubules, myxomatous stroma and low-grade nuclear cytology. Proliferation of spindle cells or foci of clear cells are also observed. Histochemically, the myxomatous stroma exhibits a positive reaction for alcian blue and colloidal iron stainings. Ultrastructurally, short microvilli are focally observed and junctional complexes are present. Recently, multiple losses of chromosomes 1, 4, 6, 8, 9, 13, 14, 15 and 22 in MTSCC have been elucidated by using comparative genomic hybridization. The prognosis of MTSCC is generally favorable, but some cases may show local recurrence or metastasis. Some cases with MTSCC seem to show overlapping histology with low-grade collecting-duct carcinoma. Therefore, further investigation will be needed to elucidate pathobiological characteristics of MTSCC.

Presence of vascular adventitial fibroblastic cells in diffuse-type gastric carcinomas.

AIM: To investigate morphological changes in the tumour vessel adventitia, particularly the distribution of vascular adventitial fibroblastic cells (VAFCs)--namely, CD34 positive fibroblastic cells just outside the vascular media--in diffuse-type gastric carcinomas. METHOD: In total, 18 surgically resected advanced typical diffuse-type gastric carcinomas and their normal tissues were examined. Immunostaining for CD34, CD31, high molecular weight caldesmon (HCD), and cytokeratin 8 (CAM5.2) was performed to detect VAFCs. VAFCs are positive for CD34 but negative for CD31, and are located just outside the vascular media (HCD positive vascular smooth muscle bundle). The areas just outside the vascular media in the whole maximum tumour cut surface were assessed, except the tumour growing edge, which was confirmed by immunostaining with CAM5.2. CD34 positive and CD31 negative cells just outside the vascular media were defined as VAFCs. RESULTS: VAFC containing vessels were seen in 17 of the 18 diffuse carcinoma tissues. Vessels lacking VAFCs were also detected in these 17 tumours. In contrast, all of the vessels lacked VAFCs in the remaining tumour. In the 18 samples of normal tissue, all of the vessels contained VAFCs. CONCLUSIONS: These results suggest that the presence of VAFCs is associated with the infiltration of diffuse scattered gastric carcinoma cells.

The disappearance of CD34-positive and alpha-smooth muscle actin-positive stromal cells associated with human intra-uterine and tubal pregnancies.

In order to elucidate the change in alpha-smooth muscle actin (ASMA)-positive and CD34-positive stromal cells associated with pregnancy, we examined endometrial and Fallopian tube tissues from 40 patients including normal endometrium (n=10), intra-uterine pregnancy (n=10), normal Fallopian tube (n=10), and tubal pregnancy (n=10), using immunohistochemistry. In normal endometrium, only a few ASMA-positive cells were focally observed. Additionally, a wide range of CD34-positive stromal cell abundance was observed. In normal Fallopian tube mucosa, a small to moderate number of both ASMA-positive and CD34-positive stromal cells was observed. Neither ASMA-positive nor CD34-positive stromal cells were observed anywhere in the decidual stroma during both intra-uterine and tubal pregnancies. Likewise, a varying abundance of ASMA-positive cells but no CD34-positive stromal cells were observed at the fetal side during both intra-uterine and tubal pregnancies. In conclusion, the disappearance of CD34-positive and ASMA-positive stromal cells may be an indicator of decidualisation induced change in the stroma during both intra-uterine and tubal pregnancies. ASMA-positive stromal cells at the fetal side associated with pregnancy may play a role in the production of villous extracellular matrix or regulation of blood flow.

Distribution and role of CD34-positive stromal cells and myofibroblasts in human normal testicular stroma.

CD34-positive stromal cells are distributed in various organs including breast, Fallopian tubes, thyroid gland, colon, pancreas, and uterine cervix. To elucidate the distribution of CD34-positive stromal cells, smooth muscle cells, and myofibroblasts in normal human testis, we examined 48 testes obtained by autopsy and operation, including five fetal, one neonatal, and 42 adult cases without evident testicular lesions, using a streptavidin-biotin immunoperoxidase technique. The expression of alpha-smooth muscle actin (ASMA), h-caldesmon, CD34, and CD31 were immunohistochemically examined in all cases. The tunica albuginea and the inner layer of seminiferous tubules in adult testis were predominantly composed of myofibroblasts. Smooth muscle cells were also scattered throughout these sites in some cases. CD34-positive stromal cells were abundant, and they formed a reticular network around the seminiferous tubules and Leydig cells as well as the outer layer of seminiferous tubules. Moreover, myofibroblasts and the CD34 reticular network were already present in the testicular stroma during fetal or neonatal development. Double immunostaining of fetal, neonatal and adult testes using ASMA and CD34 confirmed that myofibroblasts and CD34-positive stromal cells were present in the inner and outer layers of peritubular tissue, respectively. This distribution and cytological identification was also confirmed by an ultrastructural study of four cases. Finally, CD34-positive stromal cells and myofibroblasts are major components of human testicular stroma.

Lack of vascular adventitial fibroblastic cells in tumour stroma of intestinal-type and solid-type gastric carcinomas.

AIMS: To investigate the roles of vascular adventitial fibroblastic cells in tumour stroma, the distribution of vascular adventitial fibroblastic cells was studied in gastric carcinomas. METHODS: In total, 50 surgically resected gastric carcinomas (43 intestinal type, and seven solid type) and their normal tissues were examined. Vascular adventitial fibroblastic cells are positive for CD34 but negative for CD31. To differentiate vascular adventitial fibroblastic cells from vascular endothelial cells, immunostaining for CD34 and CD31 was performed. Immunostaining for high molecular weight caldesmon was also performed to recognise vascular media. RESULTS: In normal gastric tissues, CD34 positive fibroblastic cells were found just outside the vascular media, namely vascular adventitial fibroblastic cells. In contrast, all of the 43 intestinal-type and seven solid-type gastric carcinomas had no vascular adventitial fibroblastic cells in the tumour stroma. CONCLUSIONS: These results suggest that a lack of vascular adventitial fibroblastic cells is associated with tumour stroma formation in intestinal-type and solid-type gastric carcinomas.