A case of breast milk-acquired cytomegalovirus infection in an extremely low birth weight infant.

INTRODUCTION: Although breast milk is considered the optimal nutrition for infants, it is also the primary cause of postnatal cytomegalovirus (CMV) infection. Preterm infants with postnatal CMV infections are susceptible to a variety of life-threatening conditions. CASE SUMMARY: Twin male infants were delivered via emergency caesarian section at 27 weeks' gestation secondary to maternal complete uterine rupture. The Apgar scores at 1 and 5 min were 1 and 1 for the older twin (Twin A) and 0 and 3 for the younger twin (Twin B). Their birth weights were 1203 g (+ 0.65SD) and 495 g (- 3.79SD) respectively. On day 41, laboratory blood test results for Twin B showed a moderate elevation in C-reactive protein (CRP), thrombocytopenia. CMV quantitative polymerase chain reaction (qPCR) tests in Twin B's urine and blood as well as in the mother's breast milk were positive, but stored, dried umbilical cord CMV qPCR tests were negative. Twin B was diagnosed with a postnatal CMV infection secondary to infected breast milk and ganciclovir was commenced on day 52. Treatment was switched to valganciclovir at 74 days of age, but a negative CMV-DNA level in the blood was not achieved. Postnatal CMV infection in this infant led to an exacerbation of pre-existing bronchopulmonary dysplasia (BPD) and he demised at 182 days of age. CONCLUSION: Postnatal cytomegalovirus infections may lead to exacerbations of BPD. Early use of raw breast milk in preterm infants should be done with careful consideration of this potential complication.

Precise analysis of the timing performance of Cherenkov-radiator-integrated MCP-PMTs: analytical deconvolution of MCP direct interactions.

In order to achieve the ultimate goal of reducing coincidence time resolution (CTR) to 10 ps, thus enabling reconstruction-less positron emission tomography, a Cherenkov-radiator-integrated microchannel plate photomultiplier tube (CRI) reaching CTR of sub-50 ps full width at half maximum (FWHM) has been developed. However, a histogram of time differences between a pair of the CRIs shows undesirable side peaks, which are caused by gamma rays directly interacting with the micro channel plates (MCPs). Such direct interaction events are detrimental to the timing performance of the CRI. In this paper, we demonstrate an analytical method of deconvolving MCP direct interaction events from the timing histogram. Considering the information of the main and the two side peaks, the timing uncertainty caused by the MCP direct interaction events is deconvolved and the CTR of the CRI is analytically investigated. Consequently, the CTR is improved from 41.7 to 40.5 ps FWHM by the deconvolution. It means that a mixture of the Cherenkov radiator events and the MCP direct interaction events contribute to the CTR by a factor of 10 ps. The timing performance of the MCP direct interaction events are also evaluated. The CTR between the two MCPs is found to be 66.2 ps FWHM. This indicates that a photocathode-free radiation detector with high timing performance is possible. Elimination of the photocathode from the detector would make detector construction easier and more robust.

[Endobronchial hamartoma suspected lung cancer due to false positive of fluorodeoxyglucose-positron emission tomography; report of a case].

A 63-year-old male was referred to our hospital with an abnormal shadow on chest radiography and computed tomography. A transbronchial lung biopsy did not reveal any malignant lesion. A fluorodeoxyglucose-positron emission tomography (FDG-PET) was done for further examination and it revealed positive. Right upper lobectomy was perfomed for both diagnosis and treatment. The pathological diagnosis was endobronchial hamartoma The FDG-PET was false positive because of its atelectasis with obstructive pneumonia

Coincidence time resolution of 30 ps FWHM using a pair of Cherenkov-radiator-integrated MCP-PMTs.

Radiation detectors dedicated to time-of-flight positron emission tomography (PET) have been developed, and coincidence time resolution (CTR) of sub-100 ps full width at half maximum (FWHM) has been achieved by carefully optimizing scintillators and photodetectors. Achieving a CTR of 30 ps FWHM by using a pair of annihilation γ-rays would allow us to directly localize the annihilation point within an accuracy of 4.5 mm. Such direct localization can potentially eliminate the requirement of image reconstruction processes in clinical PET systems, which would have a huge impact on clinical protocols and molecular imaging. To obtain such a high CTR, researchers have investigated the use of prompt emissions such as Cherenkov radiation and hot-intra band luminescence. Although it is still challenging to achieve a CTR of 30 ps FWHM even with a Cherenkov-based detector, the experimentally measured CTR is approaching the goal. In this work, we developed a Cherenkov-radiator-integrated micro-channel plate photomultiplier tube (CRI-MCP-PMT), where there are no optical boundaries between the radiator and photocathode, and its timing performance was investigated. By removing the optical boundaries, reflections are eliminated and transmission to the photocathode is improved, resulting in high timing capability. As a result, a CTR of 30.1 ± 2.4 ps FWHM, which is equivalent to a position resolution of 4.5 ± 0.3 mm along a line of response (LOR), was obtained by using a pair of CRI-MCP-PMTs.

Evolution of prokaryotic gene order: genome rearrangements in closely related species.

Conservation of gene order in prokaryotes has become important in predicting protein function because, over the evolutionary timescale, genomes are shuffled so that local gene-order conservation reflects the functional constraints within the protein. Here, we compare closely related genomes to identify the rate with which gene order is disrupted and to infer the genes involved in the genome rearrangement.

TAP (NXF1) belongs to a multigene family of putative RNA export factors with a conserved modular architecture.

Vertebrate TAP (also called NXF1) and its yeast orthologue, Mex67p, have been implicated in the export of mRNAs from the nucleus. The TAP protein includes a noncanonical RNP-type RNA binding domain, four leucine-rich repeats, an NTF2-like domain that allows heterodimerization with p15 (also called NXT1), and a ubiquitin-associated domain that mediates the interaction with nucleoporins. Here we show that TAP belongs to an evolutionarily conserved family of proteins that has more than one member in higher eukaryotes. Not only the overall domain organization but also residues important for p15 and nucleoporin interaction are conserved in most family members. We characterize two of four human TAP homologues and show that one of them, NXF2, binds RNA, localizes to the nuclear envelope, and exhibits RNA export activity. NXF3, which does not bind RNA or localize to the nuclear rim, has no RNA export activity. Database searches revealed that although only one p15 (nxt) gene is present in the Drosophila melanogaster and Caenorhabditis elegans genomes, there is at least one additional p15 homologue (p15-2 [also called NXT2]) encoded by the human genome. Both human p15 homologues bind TAP, NXF2, and NXF3. Together, our results indicate that the TAP-p15 mRNA export pathway has diversified in higher eukaryotes compared to yeast, perhaps reflecting a greater substrate complexity.

Re-annotating the Mycoplasma pneumoniae genome sequence: adding value, function and reading frames.

Four years after the original sequence submission, we have re-annotated the genome of Mycoplasma pneumoniae to incorporate novel data. The total number of ORFss has been increased from 677 to 688 (10 new proteins were predicted in intergenic regions, two further were newly identified by mass spectrometry and one protein ORF was dismissed) and the number of RNAs from 39 to 42 genes. For 19 of the now 35 tRNAs and for six other functional RNAs the exact genome positions were re-annotated and two new tRNA(Leu) and a small 200 nt RNA were identified. Sixteen protein reading frames were extended and eight shortened. For each ORF a consistent annotation vocabulary has been introduced. Annotation reasoning, annotation categories and comparisons to other published data on M.pneumoniae functional assignments are given. Experimental evidence includes 2-dimensional gel electrophoresis in combination with mass spectrometry as well as gene expression data from this study. Compared to the original annotation, we increased the number of proteins with predicted functional features from 349 to 458. The increase includes 36 new predictions and 73 protein assignments confirmed by the published literature. Furthermore, there are 23 reductions and 30 additions with respect to the previous annotation. mRNA expression data support transcription of 184 of the functionally unassigned reading frames.

Novel flushing provoked by volatile anesthetics in Mastomys natalensis bearing a transplantable substrain of gastric carcinoid that predominantly secretes serotonin.

A tumor substrain secreting a large amount of serotonin [5-hydroxytryptamine (5-HT); CAS: 50-67-9; 3-(2-amino-ethyl)indol-5-ol] and a minute amount of histamine (CAS: 51-45-6) has been isolated from the previously established strain of transplantable gastric carcinoid of Mastomys (Praomys) natalensis secreting both histamine and 5-HT. Mastomys bearing a large growing transplant and excreting a large amount of 5-hydroxy-indoleacetic acid [(5-HIAA) CAS: 54-16-0] were associated often with reddening of the nose, lower lip, auricles, hands, and feet. Soon after the animals were anesthetized by ether or other volatile anesthetics, the tinges of red of the above-mentioned exposed parts abruptly turned bright red and rapidly spread over the neck, upper chest, and epigastric area. The reddening was transient, lasting 1.5-5 minutes, thereby fulfilling the criteria of flushing. The severity of ether-provoked flushing in tumor-bearing Mastomys paralleled the urinary excretion levels of 5-HIAA. The ether-provoked flushing was prevented completely by sc injection of either ketanserin (150 micrograms) or somatostatin (20 micrograms). The same ether-provoked flushing as found in tumor-bearing Mastomys could be reproduced in normal ones by constant infusion of 20 mg 5-HT/kg/24 hours (i.e., doses comparable to those released from a transplanted tumor) through an osmotic minipump implanted subcutaneously.

Production of serotonin by a transplantable strain of histamine-producing primary gastric carcinoid of Mastomys natalensis.

Two biochemically distinguishable transplantable tumor strains (A and B) were established from a primary gastric carcinoid of Mastomys secreting histamine alone. Strain A in the third generation acquired a new ability to produce serotonin [5-hydroxytryptamine (5-HT)] and retained endocrine activities to produce both histamine and 5-HT through the following subpassages, whereas strain B (like the primary tumor) continued to produce histamine alone. The findings were further supported by the immunohistochemical demonstration of 5-HT-containing tumor cells in strain A after generation 3 and the absence of such cells in strain B and also by the ultrastructural demonstration of tumor cells containing pleomorphic secretory granules in strain A after the third generation but not in strain B. Sixteen samples of Formalin-fixed, paraffin-embedded primary gastric carcinoids of Mastomys were stained by the same immunohistochemical method for 5-HT detection. The positively stained tumor cells were demonstrated in 4 tumor samples, though they were scantily distributed in tumor parenchyma except for 1 metastasizing tumor. 5-HT-producing tumor cells appeared through many proliferative cycles of the deranged histamine-producing cells. The endocrinologic similarity was noted between this transplantable tumor strain and a specific type of gastric carcinoid in humans, and the possible histogenesis of the latter tumor was discussed on the basis of data obtained from the present transplantation experiments.

Clinical implications of p53 autoantibodies in the sera of patients with non-small-cell lung cancer.

BACKGROUND: The presence of autoantibodies to p53 protein has been associated with the presence of p53 (also known as TP53) gene mutations in primary tumors and with poor prognosis. This study was undertaken to determine the clinical significance of p53 autoantibodies in patients with non-small-cell lung cancer (NSCLC). METHODS: We studied 188 consecutive patients with NSCLC who underwent pulmonary resection and for whom preoperative serum was available. The presence of p53 autoantibodies, detected by use of two amino-terminal and two carboxy-terminal peptides (20-30 mers) as antigens and an enzyme-linked immunosorbent assay, was related to various clinicopathologic parameters and to overexpression of p53 protein in the primary tumor. For 22 patients who had p53 autoantibodies before surgery, we also examined sera taken during postoperative follow-up. Reported P values are two-sided. RESULTS: Autoantibodies to p53 protein were detected in 38 patients. Patients with squamous cell carcinoma, those with more advanced disease (stage III-IV), and those with tumors that overexpressed p53 had a significantly higher incidence of p53 autoantibodies (P = .05,.0079, and .02, respectively). In all but one of the patients with postoperative serum samples, the antibody titer declined after surgery; however, there was no relationship between clinical course and this change in antibody titer. In addition, there was no relationship between the presence of p53 autoantibodies and overall survival in 171 patients who underwent potentially curative resection (P = .28); however, 13 patients with autoantibodies to amino-terminal peptides had a worse overall survival (P = .02). CONCLUSIONS: In NSCLC, the incidence of p53 autoantibodies is associated with histologic type, stage, and p53 overexpression--but not with patient survival. Our data do not support the clinical utility of p53 autoantibodies as diagnostic or prognostic markers in patients with NSCLC.

p53 mutations in non-small cell lung cancer in Japan: association between mutations and smoking.

The p53 gene has been implicated as a tumor suppressor gene involved in the pathogenesis of lung cancer. Our previous study revealed that the p53 gene is frequently mutated with a distinct nucleotide substitution pattern in small cell lung cancer specimens in Japanese patients. In this study, we examined 30 primary, resected non-small cell lung cancer samples in Japanese patients using complementary DNA-polymerase chain reaction and sequencing. Mutations changing the p53 coding sequence were found in 14 of 30 tumor samples (47%), while G:C to T:A transversions which are uncommon in other cancers such as colon cancer were the most frequently observed mutations, in agreement with an earlier report on non-small cell lung cancer in American patients. Furthermore, the present study shows for the first time that in univariate and multivariate analyses, the presence of p53 mutations is closely associated with lifetime cigarette consumption.

Preferential expression of c-kit protooncogene transcripts in small cell lung cancer.

As an initial step to understand rapid growth of small cell lung cancer (SCLC), a complementary DNA library prepared from a SCLC cell line was screened with viral oncogene probes encoding protein-tyrosine kinases, which are known to play an important role in regulation of cell growth. Fifteen clones hybridizing with v-fms probe were isolated, and, by partial sequence analysis, four of them were identified to be c-kit protooncogenes. Northern blot study demonstrated that most of the SCLC tumors and cell lines expressed c-kit transcripts, while non-SCLC tumors and cell lines did not. Neither amplification nor rearrangement of the c-kit gene was demonstrated in SCLC cell lines by Southern blot analysis, however. Our results suggested that c-kit expression in SCLC reflects the unique biological nature of the tumor cells different from non-SCLC and further suggested that the c-kit product may participate in autocrine or paracrine stimulation of SCLC growth.

Prognostic significance of p53 mutations and 3p deletions in primary resected non-small cell lung cancer.

We evaluated the prognostic significance of p53 mutations and an allelic loss of chromosome 3p in 71 patients with non-small cell lung cancer who underwent potentially curative resection. p53 mutations were detected in 35 cases (49%), while 3p deletions were observed in 34 of 70 informative cases (49%). The presence of the p53 mutation was associated with a shortened survival in all patients (P = 0.014 by log rank test), including those in early stages of the disease (stage I or II, n = 48) (P = 0.016 by log rank test). Multivariate analysis by the Cox proportional hazards model also revealed that p53 mutation was an independent yet unfavorable prognostic factor (P = 0.013). Patients with 3p deletion tended to have a poorer prognosis, but not to a statistically significant extent.

Distinct mutational spectrum of the p53 gene in lung cancers from Chinese women in Hong Kong.

Accumulating evidence suggests that the p53 gene is a good target for molecular epidemiological studies to search for risk factors in carcinogenic events. The lung cancer incidence for females in Hong Kong is unusually high, ranking among the highest in the world despite a low percentage with a history of smoking. To gain insights into possible etiological risk factors responsible for this high incidence, we examined p53 mutations in 35 lung cancer specimens from Chinese females living in Hong Kong and compared them with 35 matched cases from Japanese women as well as previously reported p53 mutations in the world literature. p53 mutations in exons 5-8 were present in 20 and 31% of the Hong Kong and Japanese cases, respectively. Notably, single-base deletions within runs of identical bases were observed in 3 (43%) of the 7 mutations in the Hong Kong cases, in contrast to the absence of such mutations in the controls and the extreme scarcity in the literature, suggesting that distinct environmental and/or genetic factor(s) might be involved. Although the frequent occurrence of characteristic single-base deletions could be a reflection of mutator mutations leading to inefficient mismatch repair of slipped strand mispairings, none of the lung cancer specimens exhibited such microsatellite instabilities.

Establishment of human peripheral lung epithelial cell lines (HPL1) retaining differentiated characteristics and responsiveness to epidermal growth factor, hepatocyte growth factor, and transforming growth factor beta1.

Novel human epithelial cell lines retaining characteristic features of normal peripheral airway cells were established by transfecting the SV40 large T antigen gene into primary in vitro outgrowths from normal peripheral lung specimens. These lines, designated as HPL1A to HPL1E, showed the polygonal shapes typical of epithelial cells and expressed cytokeratin in abundance. Ultrastructural examination revealed the presence of microvilli, multivesicular bodies, and multilamellar body-like structures that are characteristic of type II pneumocytes, but expression of CC1O transcripts, a highly specific marker for Clara cells, was also observed. Response to transforming growth factor beta, epidermal growth factor (EGF), and hepatocyte growth factor, all of which are thought to be important growth-regulatory molecules for cellular proliferation and developmental processes of peripheral lung, was apparent. In the HPL1A case, markedly altered cell morphology and cytoskeletal organization, potent inhibition of cell growth, and increased expression of an extracellular matrix protein were noted with transforming growth factor beta. Interestingly, both EGF and hepatocyte growth factor stimulated anchorage-dependent growth, whereas only EGF could sustain anchorage-independent proliferation. The HPL1 lines are, to our knowledge, the first series of stable epithelial lines of human peripheral lung to be described. They should be valuable for investigating various aspects of growth regulation and oncogenic processes, including the mechanisms of acquisition of anchorage independence and the interrelationships of genetic changes identified previously in lung cancers. In addition, the HPL1 lines may also prove useful for development of in vitro models for other human lung disorders as well as to elucidate the mechanisms of peripheral lung differentiation.

Chemosensitivity and radiosensitivity of small cell lung cancer cell lines studied by a newly developed 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) hybrid assay.

The 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) hybrid assay was developed by technically combining the human tumor clonogenic assay and the MTT assay to make the most of both assays. This assay was able to estimate the in vitro growth of cultured cell lines and of tumor cells in pleural effusion, suggesting the possibility of its use for assessment of chemosensitivity and radiosensitivity of fresh tumor samples. Multiple cell lines [including morphological and/or phenotypical in vitro converters and cisplatin (CDDP)-resistant lines] were established from three patients with small cell lung cancer at different stages of the disease. Chemosensitivity of these cell lines to four commonly used chemotherapeutic drugs was tested by the MTT hybrid assay. SK1 and SK3 lines were established from Patient S. K. before and after chemotherapy and radiotherapy, respectively. SK3/CDDP, a CDDP-resistant line derived from the SK3 line, was 30-fold more resistant to CDDP [50% inhibiting dose (IC50), 21.5 micrograms/ml] than the SK1 line. In Patient M. O., MOA2/CDDP, a CDDP-resistant line derived from MOA2 (an in vitro converter from the MO line), was 41-fold more resistant to CDDP (IC50, 37 micrograms/ml) than the parent MO line. From Patient T. M., TM1 and TM2 lines were established before and after chemotherapy, respectively. The latter showed 6-fold more resistance to CDDP than the former. Chemosensitivity of these lines to three other drugs, 4-hydroperoxycyclophosphamide, Adriamycin, and etoposide, suggested cross-resistance between CDDP and 4-hydroperoxycyclophosphamide. Radiosensitivity study was also carried out with the MTT hybrid assay. The MOA2 line was more resistant [Do, 3.0 Gy; extrapolation number (n), 4.0] than the parental MO line (Do, 1.6 Gy; n, 2.1). There was no clear difference in radiosensitivity between the cell lines established before and after radiation therapy in Patient S. K.

Three distinct regions involved in 3p deletion in human lung cancer.

The 3p deletion was first noted by cytogenetic analysis and was later confirmed by several independent studies using restriction fragment length polymorphism (RFLP) probes. As an initial step towards positional cloning (reverse genetics) of the tumor-suppressor gene(s) on 3p, a detailed analysis of the minimum deleted region(s) on 3p was performed with 13 RFLP probes and 48 paired human lung cancer samples. All nine small-cell lung cancer cases (100%) and 31 of 39 non-small-cell lung cancer cases (79%) showed allelic loss at one or more loci mapped on 3p. We show here that three distinct regions on 3p appear to be frequently deleted in lung cancer. These regions include 3p25, 3p21.3 and 3p14-cen. The present study should warrant future work focusing on these chromosomal regions on 3p, and may ultimately lead to the isolation of tumor-suppressor genes involved in the pathogenesis of lung cancer.

Prognostic significance of abnormal p53 accumulation in primary, resected non-small-cell lung cancers.

PURPOSE: This study was conducted to evaluate the prognostic significance of p53 abnormalities in primary, resected non-small-cell lung cancer (NSCLC). PATIENTS AND METHODS: Methodologic validation of immunohistologic detection of p53 abnormalities in routine pathology sections was assessed using 31 lung cancer specimens for which p53 gene status was known from our previous molecular biologic studies. Applying the optimized cutoff value, we evaluated the prognostic significance of p53 abnormalities in an independent cohort of 208 NSCLC patients with complete follow-up data, whose resections were consecutively performed between January 1984 and December 1988. RESULTS: Immunohistologic detection of p53 abnormalities appeared to be reliable and showed approximately 90% concordance with the p53 gene status. Using the selected cutoff value of 10%, 46% of 208 NSCLCs showed p53 abnormalities. There was no relationship between p53 abnormalities and clinical outcome in the entire cohort, which represented all histologic subtypes of NSCLC (P = .58). Based on the reasoning that the influence of p53 abnormalities may have been obscured by distinct biologic roles depending on histologic subtypes, we also separately analyzed subsets of patients with adenocarcinomas (n = 100) and with squamous cell carcinomas (n = 88) and found that it may be a useful prognosticator only in adenocarcinoma patients (P = .04). CONCLUSION: p53 abnormalities are not a significant prognostic factor in primary, resected NSCLC when all histologic subtypes are combined, but may be a useful prognosticator for adenocarcinomas. Additional studies are warranted for further evaluation, specifically of adenocarcinomas.

High frequency of clonally related tumors in cases of multiple synchronous lung cancers as revealed by molecular diagnosis.

In patients with multiple synchronous lung tumors, discrimination of multicentric lung cancers from intrapulmonary metastasis is important for treatment decision, but this is sometimes difficult. The aim of this study was to retrospectively distinguish multicentric lung cancers from intrapulmonary metastases in 14 such cases by loss of heterozygosity (LOH) and p53 mutational status. DNA was extracted from microdissected tumor cells in paraffin-embedded archival tissue, and 3p14.2, 3p21, 3p25, 9p21, and 18q21.1 were investigated for LOH. Exons 5-8 of the p53 gene were examined for mutations by the PCR, followed by single-strand conformation polymorphism analysis and DNA sequencing. For cases with the same LOH pattern, we calculated a clonality index, the probability of the given LOH pattern when these tumors were hypothesized to be independent in origin. Eleven of 14 cases (79%) were thus diagnosed as having pulmonary metastasis and only one case as having genuinely multicentric lung cancers. Two cases presented difficulty in diagnosis. In several cases, the LOH patterns conflicted with p53 mutation patterns, suggesting that clonal evolution is directly affected by certain genetic changes. The combination of p53 with LOH helped increase both the sensitivity and specificity of the assay.