Association of adenocarcinoma of the lung with cigarette smoking by grade of differentiation and subtype.

A hospital-based case-control study was carried out in order to evaluate the risk of adenocarcinoma of the lung associated with cigarette smoking according to grade of differentiation and subtype. The cases studied were 238 patients with adenocarcinomas of the lung (158 males and 80 females) that were surgically resected at the Center for Adult Diseases, Osaka. For each case, 2 controls were chosen at the same hospital from outpatients who had not been diagnosed as having smoking-related diseases, matched by sex, age, and year of first visit. When the male cases with adenocarcinoma were classified according to the grade of differentiation, the odds ratios (ORs) associated with exsmokers and current smokers were: 1.0, 2.1 for well-differentiated; 4.1, 7.7 for moderately differentiated; and 8.5, 7.9 for poorly differentiated adenocarcinoma. The OR associated with current smokers for poorly and moderately differentiated adenocarcinoma combined was significantly higher than that for well differentiated adenocarcinoma. Approximately the same pattern of ORs was observed in females. For poorly and moderately differentiated adenocarcinoma, a significant dose-response relationship was observed in males. Comparison between the ORs for papillary type and tubular type showed no difference.

Reduced motility related protein-1 (MRP-1/CD9) gene expression as a factor of poor prognosis in non-small cell lung cancer.

Motility related protein-1 (MRP-1) is a transmembrane glycoprotein that is identical to the CD9 antigen. In previous studies, we showed that various types of cultured tumor cells transfected with MRP-1/CD9 cDNA have low motility and diminished metastatic potential to the lung. More recently we used immunohistochemical procedures, immunoblotting, and reverse transcription-PCR to demonstrate that the level of MRP-1/CD9 expression was inversely related to the clinical stage of a given carcinoma of the breast. In addition, we found that the primary tumors of almost 50% of the patients had higher MRP-1/CD9 levels than their respective metastatic lymph nodes. In consideration of these findings, we have now applied reverse transcription-PCR to determine MRP-1/CD9 gene expression in lung cancer. We analyzed tumor tissues of 109 patients: 49 tumors were stage I; 15 were stage II; and 45 were stage III. We found that 67 patients had MRP-1/CD9-positive tumors, and that gene expression was reduced in the tumors of the remaining 42 individuals. The overall rate of survival was strikingly higher among patients with positive tumors than in those whose tumors had reduced gene expression (62.3 versus 34.9%; P < 0.001). This also pertained to patients with adenocarcinomas of the lung (55.4 versus 26.0%; P < 0.001). Multivariate analysis with the Cox regression model indicated that MRP-1/CD9 positivity correlated better with overall survival rate than did other variables, except lymph node status. Our data suggest that low MRP-1/CD9 expression by tumors of the lung may be associated with poor prognosis. It is conceivable that testing for MRP-1/CD9 may identify node-negative lung cancer patients and patients with adenocarcinomas who are at high risk for early disease recurrence.

Expression of alpha-amylase in human lung cancers.

Thirty-three human lung tumors were studied for the expression of alpha-amylase by immunohistochemical and Northern blot analyses. Twenty of them were adenocarcinomas, among which 17 proved to be adequate for mRNA analyses and were, except for two, amylase mRNA producers. Seven were squamous cell carcinomas, none of which produced amylase. The remaining six consisted of two undifferentiated small cell carcinomas, and one each of undifferentiated large cell carcinoma, carcinoid tumor, mucoepidermoid carcinoma, and metastatic lung cancer; the mucoepidermoid carcinoma proved to be an amylase producer. These observations strongly suggest that among lung cancers, the production of alpha-amylase is a property commonly associated with adenocarcinomas and can be used for distinguishing cell types. Histogenesis and carcinogenesis in lung cells are discussed in connection with the cells that produce amylase.

Retention of acyl groups in LM cell fibroblasts with altered phospholipid composition.

Incorporation and loss of membrane lipids were measured in murine fibroblasts grown in suspension culture with [2-14C]acetate in a chemically defined medium devoid of serum or lipid. Greater than 98 and 94% of the 14C present in the phospholipids and neutral glycerides respectively was found in the fatty acyl groups. During a 3-day growth period LM fibroblasts lost 65% of their 14C-labelled acyl groups from the phospholipids and 36% from the neutral lipids. Selective retention of 14C in individual isolated phospholipid and neutral lipid species occurred only in phosphatidylethanolamine. Retention of acyl groups in the phospholipids or neutral glycerides was not a property of LM suspension cultured fibroblasts. Supplementation of the LM cells with choline analogues, such as N,N'-dimethylethanolamine, N-monomethylethanolamine, or ethanolamine instead of choline, decreased the loss of 14C-labelled fatty acids from phospholipid and neutral lipid. Ethanolamine supplementation resulted in increased loss of 14C from desmesterol as well as decreased incorporation of 14C into desmosterol. Thus, polar head group manipulation affected acyl group composition, acyl group retention, and sterol metabolism in suspension cultured LM cells.

Behavior of vesicular stomatitis virus glycoprotein in mouse LM cells with modified membrane-phospholipids.

LM cells in which the membrane phospholipids had been modified with choline analogues were infected with vesicular stomatitis virus. The choline analogues tested were choline, N,N'-dimethylethanolamine, N-monomethylethanolamine and ethanolamine. These modifications per se did not affect the syntheses of individual viral proteins. The viral glycoprotein was detected in the plasma membranes of all the modified cells by pronase digestion in pulse-chase experiments, but the amount of glycoprotein susceptible to proteolysis varied, decreasing in these modified cells in the following order: N,N'-dimethylethanolamine- greater than choline- greater than N-monomethylethanolamine- greater than ethanolamine-treated cells. After a 4-h chase, glycoprotein was mainly distributed in the plasma membranes of cells modified with N,N'-dimethylethanolamine, whereas it was found in both the microsomes and plasma membranes of cells modified with other analogues. Fairly large amounts of glycoprotein were also found in the soluble fraction of ethanolamine-treated cells, but not in that of choline- or N,N'-dimethylethanolamine-treated cells. More precise experiments on the behaviour of glycoprotein with a short period of chase strongly suggested that migration of glycoprotein from the microsomes to the plasma membranes was fastest in cells modified with N,N'-dimethylethanolamine and slowest in cells modified with ethanolamine. Membrane lipid modifications also resulted in release of different numbers of progeny virions from the cells, release of virions from the cells decreasing in the following order: N,N'-dimethylethanolamine- greater than choline- greather N-monomethylethanolamine- greater than ethanolamine-treated cells. These results indicate that modification of membrane phospholipids influences not only the insertion of glycoprotein into the microsomes and its migration to the plasma membranes, but also the production of progeny virions.

Synthesis of acyl phosphatidylglycerol from phosphatidylglycerol in Escherichia coli K-12. Evidence for the participation of detergent-resistant phospholipase A and heat-labile membrane-bound factor(s).

The conversion of phosphatidylglycerol to acyl phosphatidylglycerol by extracts of Escherichia coli K-12 strains was examined under various conditions. The maximum rate of conversion was observed at pH 7.2 in the presence of 50% (v/v) diethyl ether and 10 mM CaCl2. This conversion was found to involve two sequential reactions: (1) The formation of 2-acyl glycerophosphoglycerol and 2-acyl glycerophosphoethanolamine from phosphatidylglycerol and phosphatidylethanolamine, respectively, by detergent-resistant phospholipase A in the presence of Ca2+ and (2) transfer of the acyl group of 2-acyl lysophospholipid to phosphatidylglycerol by a heat-labile factor(s) in the presence of diethyl ether. Neither fatty, acid acyl-CoA nor 1-acyl lysophospholipid could act as an acyl donor for phosphatidylglycerol. The heat-labile factor(s) was found in both the inner membrane and supernatant fractions.

Manipulation of fatty acid composition of membrane phospholipid and its effects on cell growth in mouse LM cells.

Fatty acid composition of the phospholipids of mouse LM cells grown in suspension culture in serum-free chemically defined medium was modified by supplementing the medium with various fatty acids bound to bovine serum albumin. Following supplementation with saturated fatty acids of longer than 15 carbons (100 micron) profound inhibition of cell growth occurred; this inhibitory effect was completely abolished when unsaturated fatty acids were added at the same concentration. Supplementing with unsaturated fatty acids such as linoleic acid, linolenic acid or arachidonic acid had no effect on the cell growth. Fatty acid composition of membrane phospholipids could be manipulated by addition of different fatty acids. The normal percentage of unsaturated fatty acids in LM cell membrane phospholipids (63%) was reduced to 35--41% following incorporation of saturated fatty acids longer than 15 carbon atoms and increased to 72--82% after addition of unsaturated fatty acids. A good correlation was found between the unsaturated fatty acid content of membrane phospholipids and cell growth. When incorporated saturated fatty acids reduced the percentage of unsaturated fatty acids in membrane phospholipids to less than 50%, severe inhibition of the cell growth was found. Simultaneous addition of an unsaturated fatty acid completely abolished this effect of saturated fatty acids. The results suggest that maintenance of membrane fluidity by unsaturated fatty acids in membrane phospholipids is critical to membrane integrity and cell growth.

Reduced integrin alpha3 expression as a factor of poor prognosis of patients with adenocarcinoma of the lung.

PURPOSE: We investigated the possible association between integrin alpha3 and motility-related protein (MRP-1), cluster of differentiation antigen 9 (CD9) gene expression in non-small-cell lung cancer (NSCLC) and evaluated the prognostic significance of integrin alpha3 expression. PATIENTS AND METHODS: We performed a retrospective study of integrin alpha3 and MRP-1/CD9 expression in resected tumor tissues from 151 NSCLC patients using quantitative reverse-transcriptase polymerase chain reaction (RT-PCR) and immunohistochemistry. RESULTS: The ratio of integrin alpha3/beta-actin expression ranged from 0 to 5.87 (mean was 0.80; median, 0.70). Using the cutoff value of 0.7, there were 78 (52%) integrin alpha3-positive tumors and 73 (48%) tumors with reduced integrin alpha3 expression. The immunohistochemical results agreed well with those of the RT-PCR assays, and 88% had no discrepancy. In case of discrepancy, the results of RT-PCR were used in specimen classification. Integrin alpha3 gene expression was independent from MRP-1/CD9 gene expression. No significant association was found between integrin alpha3 expression and the patients' clinical characteristics. The overall survival rate of patients with integrin alpha3-positive NSCLCs was only slightly better than that of individuals whose tumors had reduced integrin alpha3 expression (55.9% v 47.1%; P = .085). By comparison, the overall survival rate of patients with integrin alpha3-positive adenocarcinomas was strikingly greater than in those whose tumors had reduced gene expression (54.4% v 35.2%; P = .004). Multivariate analysis with the Cox regression model of NSCLC and adenocarcinoma indicated that integrin alpha3 expression correlated better (P = .0188 and P = .0008, respectively) with the overall survival rate than other variables, except lymph node status. CONCLUSION: No significant association was found between integrin alpha3 and MRP-1/CD9 gene expression in lung cancer. However, reduced integrin alpha3 expression is a poor prognosis factor in patients with adenocarcinomas.

Pretreatment of chick embryos with estrogen in ovo prevents mullerian duct regression in organ culture.

Diethylstilbestrol (DES) feminizes chick embryos in ovo, preventing regression of the Mullerian ducts (MDs). The feminized testis, however, despite its ovarian-like appearance, continues to produce Mullerian inhibiting substance (MIS). This study was designed to test whether exogenous DES preserved the MD against endogenous MIS by affecting the target organ (feminization). DES (0.1 mg) was injected into the air sacs of 5-day-old chick embryos. Testes were obtained from 15-day-old chick embryos and MDs from 8-day-old chick embryos and combined in organ culture for 72 h. MD regression was examined macroscopically and microscopically after 3 days in vitro in medium without DES (group A: n = 155) and with 10(-6) M DES (group B: n = 169). Four types of coculture were set up as follows: 1) Control testis with control MD; 2) DES-treated testis with DES-treated MD; 3) Control testis with DES-treated MD; 4) DES-treated testis with control MD. In medium without DES (group A), MD regression was inhibited significantly when the duct was pretreated with DES [2) and 3) vs. 1) and 4)]. The pretreated testis produced slightly less regression [4) vs. 1)], but this was not significant. Previous studies in ovo had suggested that estrogen prevented MD regression directly as well as causing feminization of the testis, because MIS secretion was not inhibited from the feminized gonad. These results show that pretreatment of the MD with DES blocks regression in vitro by a normal testis, confirming that the primary site of action of estrogen is on the duct itself.

Prognostic significance of pS2 protein expression in pulmonary adenocarcinoma.

In the present study, pS2 protein expression in pulmonary adenocarcinoma was investigated on paraffin-embedded sections obtained from 170 patients. 28 (16%) patients showed varying degrees of pS2 protein expression in the cytoplasm of tumour cells, as detected by immunohistochemical staining with anti-pS2 protein antibody. There was a significant association between pS2 protein expression and larger tumour size, and the acinar or bronchiolo-alveolar subtype. However, no significant correlations between pS2 protein status and the other clinicopathological factors, i.e. T-factor, N-factor, stage and histological differentiation, were shown. In contrast to breast cancer, patients with pS2-positive pulmonary adenocarcinomas had a significantly worse prognosis than those with pS2-negative pulmonary adenocarcinomas; this was true for stage I patients, as well as for all patients. Multivariate analysis showed that pS2 protein expression was a discriminating variable in overall survival. These findings suggest that pS2 protein status is a possible prognostic indicator in pulmonary adenocarcinoma.

An in vitro chemosensitivity test for solid human tumors using collagen gel droplet embedded cultures.

In vitro chemosensitivity testing using a collagen gel droplet embedded culture drug sensitivity test (CD-DST), was conducted with several types of solid cancer. The overall evaluable rate was 80% (443/554), including 76% for lung (n=243), 78% for breast (n=110), 87% for gastric (n=62), 83% for colorectal (n=107) cancers and 88% for 32 metastatic brain tumors. The in vitro sensitivity of breast, gastric and colorectal cancers to mitomycin C (MMC), cisplatin (CDDP), 5-fluorouracil (5-FU) and doxorubicin (DXR) was similar to the efficacy rates reported for each drug. This was also observed with lung cancer, the sensitivity of which to MMC, CDDP, vindesine (VDS) and etoposide (VP-16) was similar to the clinical efficacy. The clinical response to chemotherapy was compared with the results of in vitro chemosensitivity testing in Il patients: the clinical correlation was 91%, with a 80% true positive and 100% true negative rate. These results suggest that the CD-DST may be clinically useful by allowing the prediction of clinical response in various solid cancers.

Lymphoepithelioma-like carcinoma of the lung: analysis of two cases for Epstein-Barr virus infection.

Lymphoepithelioma-like carcinoma, which is an uncommon histological type of epithelial tumor, has been described as being closely associated with Epstein-Barr virus (EBV) infection in organs other than the lung. Recently, we experienced two surgically resected cases of pulmonary tumors mimicking lymphoepithelioma-like carcinoma. Both cases contained EBV DNA genomes as shown by polymerase chain reaction (PCR) using EBV DNA-specific primers, one positive for EBV DNA in virtually all cancer cells, and the other showing positive hybridization in a small number of cancer cells by in situ hybridization (ISH) using digoxigenin-labeled olignucletide probes for each of EBV DNA for EBV DNA. EBV-encoded RNA-1 (EBER-1) was typically detected in one case. These results are highly suggestive of EBV-associated tumors in one of the current cases, although in the other case, no such close association was determined. It seems that lymphoepithelioma-like pulmonary carcinoma, which seems extremely unusual, may be closely associated with EBV infection in tumorigenesis.

Intrathoracic chemothermotherapy following panpleuropneumonectomy for pleural dissemination of invasive thymoma.

We report a case of pleural dissemination of invasive thymoma, which was successfully treated with intrathoracic chemothermotherapy following panpleuropneumonectomy. Intrathoracic chemothermotherapy in combination with surgery may be a hopeful adjuvant treatment to control pleural disseminated lesions of invasive thymoma.

Intentional limited resection for selected patients with T1 N0 M0 non-small-cell lung cancer: a single-institution study.

OBJECTIVES: To comparatively evaluate lobectomy and limited resection for T1 N0 M0 non-small-cell lung cancer, we reviewed case series with concurrent nonrandomized controls. METHODS: Limited resection with curative intent was performed for 63 patients with T1 N0 M0 non-small-cell lung cancer over a 10-year period. These 63 patients included 46 patients who underwent a segmentectomy as an intentional limited resection. These patients had good pulmonary function and could tolerate a lobectomy in the management of their disease. The other 17 patients underwent wedge resection or segmentectomy as a compromised limited resection because they had poor pulmonary reserve or other limiting factors and could withstand a thoracotomy but could not tolerate a lobectomy in the management of their disease. RESULTS: The 5-year survival was 93% in the intentional resection group. The survival curve for this group was not different from that for 77 patients who underwent the standard operation (lobectomy plus complete mediastinal lymph node dissection) for T1 N0 M0 non-small-cell lung cancer during the same period. The frequency of local/regional recurrence in the intentional resection group was 8.7% (4/46); the recurrence in three patients was situated in the mediastinum. According to multivariate analysis, limited resection was not associated with poor survival. CONCLUSION: Segmentectomy with regional lymph node dissection, including the mediastinum, should be considered as an acceptable alternative treatment for selected patients with T1 N0 M0 disease.

Surgical management of lung metastases. Usefulness of resection with the neodymium:yttrium-aluminum-garnet laser with median sternotomy.

Between May 1969 and September 1989, 677 metastatic lesions were resected during 107 operations in 100 patients with pulmonary metastases from various primary sites at the Center for Adult Diseases, Osaka, Japan. Of those patients, 65 underwent conventional lateral thoracotomy, and 35 patients had median sternotomy. No significant difference existed in actuarial survival after the first operation to remove the metastases between the two patients groups. Furthermore, local excision of 418 lesions was performed in 25 patients with the neodymium:yttrium-aluminum-garnet (Nd:YAG) laser. Of those, 18 patients had undergone a one-stage operation for bilateral lesions through a median sternotomy approach. Although our study was not randomized, survival of the 25 patients treated with the Nd:YAG laser tends to be longer than survival of the 75 patients for whom the Nd:YAG laser was not used. We concluded that aggressive excision and evaporation of multiple lung metastases with the Nd:YAG laser under median sternotomy is a safe and promising variation in technique and that this approach will expand the scope of surgical indications for metastatic lung tumors. For a clearer demonstration of the influence of differences in surgical techniques on long-term survival it is necessary to conduct randomized prospective studies of the surgical techniques.

Cystic mucinous adenocarcinoma of the lung. Two cases of cystic variant of mucus-producing lung adenocarcinoma.

Two previously unreported cases of mucus-producing lung adenocarcinoma are presented as uncommon tumors, which are clinicopathologically different from other histologic types of lung adenocarcinoma. The tumors, showing apparently rapid development on chest roentgenograms, were tightly packed with copious mucus and resembled cystic lesions. Because they contained very few cancer cells, and these were only at the periphery, it was impossible to diagnose malignant neoplasms preoperatively through cytologic examination. The present tumors, which we described as cystic mucinous adenocarcinoma, are considered to be a cystic variant of mucus-producing lung adenocarcinoma that expands grossly by storing mucus.

p53 immunostaining in combined type small cell lung cancer. Brief report.

Abnormalities of the p53 oncogene in lung cancer have recently been reported to be more frequent in small cell lung cancer (SCLC) than in non-small cell lung cancer (non-SCLC), but their status in combined type SCLC is as yet unknown. In this study, immunohistochemical analysis using a polyclonal antibody against p53 protein was performed in 12 surgically resected specimens of combined type SCLC. Immunoreactivity of the p53 protein was found in 5 (42%) of the 12 cases, and the immunostaining pattern of the p53 protein in areas of the non-small cell carcinoma type was the same as in those of the small cell carcinoma type. Thus, it seems that the incidence of p53 abnormalities in combined type SCLC is slightly lower than in ordinary type SCLC. It is also suggested that abnormalities of the p53 oncogene in this histological type may not be a specific event related to the morphological difference between small cell carcinoma and non-small cell carcinoma.

Nature of Escherichia coli mutants deficient in detergent-resistant and/or detergent-sensitive phospholipase A.

1. Escherichia coli K-12 mutants deficient in detergent-resistant (DR) and detergent-sensitive (DS) phospholipases A and deficient in DS phospholipase A were isolated. 2. The growth, compositions of phospholipids and fatty acids and turnover of phospholipids of three mutants (DR-, DS-, DR-DS-) were compared with those of their parent (DR- was isolated in a previous study). 3. Autodegradations of membrane phospholipids of 18,000 X g supernatants and precipitates of the homogenates of these three mutants were also compared with those of the parent, and the effects of various detergents and organic solvents on these activities were examined. 4. We could not identify any significant physiological role for DR or DS phospholipase A.

Cathepsin B expression in tumour cells and laminin distribution in pulmonary adenocarcinoma.

AIMS: To determine the correlation between cathepsin B expression and laminin distribution in pulmonary adenocarcinoma tissue. METHODS: The distribution of cathepsin B and laminin was examined in 28 formalin fixed, paraffin wax embedded specimens of pulmonary adenocarcinoma tissue, using a double immunostaining technique with commercially available antibodies to cathepsin B and laminin, respectively. RESULTS: Tumour cells in 23 (82%) cases reacted to cathepsin B: 13 cases were weakly positive and 10 were strongly positive. Laminin in tumour associated basement membrane produced various staining patterns: two cases had an almost continuous distribution of laminin in tumour associated basement membrane in the tumour tissues, while a moderately discontinuous laminin distribution pattern was found in 12 cases, and a highly fragmented pattern was found in 14 cases. The degree of cathepsin B expression in tumour cells was significantly correlated with the break up of laminin staining. In some cases a discontinuous pattern of tumour associated laminin was frequently observed adjacent to cathepsin B positive tumour cell nests. CONCLUSIONS: Considering that cathepsin B has the capacity to degrade basement membrane components, including laminin, the inverse correlation shown in this study between the increase in cathepsin B expression by tumour cells and the diminution of laminin in tumour associated basement membrane could reflect local progression and spread by pulmonary adenocarcinoma.

Bronchial neurofibrosarcoma.

A 56-year-old woman was seen with the clinical features of collapse of the right lower lobe. Intrabronchial extension of a tumor was demonstrated endoscopically. Sleeve bilobectomy was performed, and a diagnosis of bronchial neurofibrosarcoma was confirmed by light and electron microscopic and immunohistochemical studies.