Clonality analysis of different histological components in combined small cell and non-small cell carcinoma of the lung

Combined small cell and non-small cell carcinoma is relatively rare in the lung. Examination of the clonal relationship of different components in this type of tumor may give a clue to the rarity. We retrieved 6 such tumors; all 6 had small cell carcinoma and adenocarcinoma components, and 3 had an additional squamous cell carcinoma component. We examined the point mutations in the p53 gene and allelic loss (ie, the loss of heterozygosity [LOH] pattern) of chromosome 3p in each component. p53 mutations were detected in the small cell carcinoma component of 5 tumors and in the non-small cell carcinoma components of 2 tumors. In 1 case, the squamous cell carcinoma component had a p53 mutation locus identical to that in the small cell carcinoma component, but in the other case, the adenocarcinoma component had a different mutation than that in the small cell carcinoma component. Chromosome 3p LOH loci in the squamous cell carcinoma component were present in the small cell carcinoma component in all 3 cases, but some LOH loci were not identical in the small cell carcinoma and adenocarcinoma components in 3 cases. These results suggest that the small cell and squamous cell carcinoma components of combined small cell lung carcinomas have an intimate clonal relationship. On the other hand, the adenocarcinoma component often may be derived from a separate clone or, more likely, undergo a progressive process separate from the squamous cell-small cell carcinoma beginning in a very early stage, that is, before the appearance of p53 and chromosome 3p abnormalities. This tumorigenesis process may explain the relative rarity of combined small cell and non-small cell carcinoma, which occurs primarily in the peripheral lung, an infrequent site of squamous cell carcinoma.     

In vivo study of the effects of cryopreservation on heart valve xenotransplantation.

BACKGROUND: Recent reports have suggested that cryopreservation reduces the immunogenicity of donor tissue. The immunomodulation by cryopreservation might influence on the tissue durability after xenotransplantation. We investigated the in vivo morphologic changes in cryopreserved xenograft (CXG) heart valves. MATERIAL AND METHOD: We transplanted a fresh (fresh xenograft; FXG) and a cryopreserved (CXG) porcine aortic root and a cryopreserved canine (cryopreserved allograft; CAG) aortic root into the abdominal aorta of a dog without any immunosuppressive agents. Explanted grafts on the 21st to 49th days after implantation were analyzed morphologically with light microscopy using some special stains, immunohistochemical analysis, and scanning electron microscopy (SEM). RESULT: Light microscopy showed the absence of smooth muscle cells in the media of the aorta in any group after transplantation. FXG valves did not maintain any cellularity after transplantation. CXG valves contained cellular infiltration in themselves. CAG valves contained numerous fibroblasts, which showed the maintenance of tissue integrity without allowing cellular infiltration. The structure of elastic fibers was well maintained, even in the part of CXG valve with cellular infiltration. Immunohistochemical studies documented the infiltration of T lymphocytes in CXG valves that were labeled by anti-CD3 antibodies. SEM demonstrated that no endothelia were seen on the surface of the valves in any group after transplantation. CONCLUSION: We concluded that the cryopreservation method might provide an immunomodulation of xenogeneic heart valves for transplantation.     

Melanin pigmentation in mammalian skin and its hormonal regulation

Cutaneous melanin pigment plays a critical role in camouflage, mimicry, social communication, and protection against harmful effects of solar radiation. Melanogenesis is under complex regulatory control by multiple agents interacting via pathways activated by receptor-dependent and -independent mechanisms, in hormonal, auto-, para-, or intracrine fashion. Because of the multidirectional nature and heterogeneous character of the melanogenesis modifying agents, its controlling factors are not organized into simple linear sequences, but they interphase instead in a multidimensional network, with extensive functional overlapping with connections arranged both in series and in parallel. The most important positive regulator of melanogenesis is the MC1 receptor with its ligands melanocortins and ACTH, whereas among the negative regulators agouti protein stands out, determining intensity of melanogenesis and also the type of melanin synthesized. Within the context of the skin as a stress organ, melanogenic activity serves as a unique molecular sensor and transducer of noxious signals and as regulator of local homeostasis. In keeping with these multiple roles, melanogenesis is controlled by a highly structured system, active since early embryogenesis and capable of superselective functional regulation that may reach down to the cellular level represented by single melanocytes. Indeed, the significance of melanogenesis extends beyond the mere assignment of a color trait.     

Selective expression of L-serine synthetic enzyme 3PGDH in schwann cells, perineuronal glia, and endoneurial fibroblasts along rat sciatic nerves and its upregulation after crush injury

Non-essential amino acid L-serine functions as a highly potent, glia-derived neurotrophic factor, because it is a precursor for syntheses of proteins, other amino acids, membrane lipids, and nucleotides, and also because its biosynthetic enzyme 3-phosphoglycerate dehydrogenase (3PGDH) is preferentially expressed in particular glial cells within the brain. Here we pursued 3PGDH expression in peripheral nerves and its change after crush injury. In the pathway of rat sciatic nerves, 3PGDH was selectively expressed in non-neuronal elements: Schwann sheaths and endoneurial fibroblasts in sciatic nerves, satellite cells in dorsal root ganglia, and astrocytes and oligodendrocytes in the spinal ventral horn. In contrast, 3PGDH was immunonegative in axons, somata of spinal motoneurons and ganglion cells, and endoneurial macrophages. One week after crush injury, 3PGDH was upregulated in the distal segment of injured nerves, where 3PGDH was intensified in activated Schwann cells and fibroblasts. 3PGDH was still negative in activated macrophages, which were instead associated or surrounded by activated Schwann cells with intensified 3PGDH. These results suggest that in the peripheral nervous system, these non-neuronal cells synthesize and may supply L-serine to satisfy metabolic demands for maintenance and regeneration of peripheral nerves and for proliferation and activation of macrophages upon nerve injury.     

Effects of added bioactive glass on the setting and mechanical properties of resin-modified glass ionomer cement

In this study, the effects of added bioactive glass on the basic setting properties of a commercially available resin-modified glass ionomer cement were investigated with respect to setting time, mechanical strength, and setting mechanism. It was found to be clinically acceptable whether the setting time was extended or shortened depending on the type of bioactive glass added. The compressive strength of the set cement containing the bioactive glass decreased and was much higher when compared with the conventional type glass ionomer cement containing bioactive glass. The Fourier-transform infrared and 13C CP/MAS-NMR spectroscopies revealed that the extent of the acid-base reaction was larger in the cements containing bioactive glass than in the commercial resin-modified glass ionomer cement because of its high basicity in the bioactive glass. The 27Al MAS-NMR showed that crosslinking of the carboxylates in the polymeric acid by Al proceeded less in the cement containing the bioactive glass.     

Identification of p46 Shc expressed in the nuclei of hepatocytes with high proliferating activity: Study of regenerating rat liver

The adaptor molecule Shc is a proto-oncogene product, and it is known to be associated with cell proliferation. However, the role of Shc in the proliferation and regeneration of hepatocytes remains unknown. In the present study, we report that p46 Shc is specifically expressed in the nuclei of proliferative (or regenerative) hepatocytes, suggesting that p46 Shc protein plays a role in hepatocellular proliferation. The expression of Shc was analyzed in liver tissue after partial hepatectomy (PH) or sham operation in Wistar rats by using immunohistochemistry and/or Western blot analysis. In addition, the expression of various cell cycle-related proteins, such as Cdk4, cyclin D1, PCNA, and Cdk1 was analyzed in the tissues of regenerating rat liver. Furthermore, the tyrosine phosphorylation of Shc was studied in liver tissue after PH or sham operation by immunoprecipitation using a monoclonal phosphotyrosine antibody. Although the protein levels of p52 Shc were unchanged in liver tissues after PH or sham operation, tyrosine phosphorylation was detected only in the regenerating rat liver after PH. The levels of p46 Shc protein were markedly increased in liver tissues during the liver regenerative process. In contrast, p66 Shc was not detected in the liver tissues after PH or sham operation. Western blotting and immunohistochemistry showed that the main location of p46 Shc was in the nuclei of proliferating hepatocytes after PH. These data suggest that p46 Shc expressed in hepatocellular nuclei may be closely related to the proliferation of hepatocytes. Therefore, it is suggested that p46 Shc expressed in hepatocellular nuclei may be a useful marker for detecting hepatocytes with high proliferative activity.     

Characterization of DNA sequences constituting the terminal heterochromatin of the chicken Z chromosome

Two clones, pCZTH5-8 and pCZTH12-8, were isolated from a female chicken genomic library by screening with sequences obtained from genomic libraries which had been constructed from a terminal region of a single Z chromosome of chicken utilizing laser microbeam irradiation and PCR amplification. Fluorescencein situ hybridization to the mitotic Z chromosome and the lampbrush ZW bivalent of chicken demonstrated that both the cloned sequences are located in the heterochromatic region of the Z chromosome at the end opposite to the pairing region with the W chromosome. The sequences pCZTH5-8 and pCZTH12-8 are distributed widely on both the telomeric bow-like loops (TBL) and the region I (short loops region) of the Z lampbrush chromosome. These clones, pCZTH12-8 particularly notably, hybridized also to the TBLs of lampbrush bivalents 1–4 of chicken. Both sequence are transcribed in the lampbrush stage oocytes on the Z chromosome and on other macrobivalents. The subfragment of pCZTH5-8 which hybridizes to the TBLs and the insert of pCZTH12-8 contain regions that are closely similar in sequence. The pCZTH5-8 sequence has no internal repeats and may be part of the 24-kb macrosatellite repeating unit that is evident afterNhel digestion of the genomic DNA. A cloned 24-kb unit, pFN-1, does not show significant DNA curvature, but cytosines of its CpG dinucleotides may be highly methylatedin vivo. This contrasts with the repeat sequences of the W heterochromatin which not only have highly methylated CpG but are also strongly curved. The 24-kb unit is repeated about 830 times in the diploid genome of a female chicken, suggesting that nearly the entire terminal heterochromatin on the Z chromosome consists of this macrosatellite family. Sequences of the greater part of the pCZTH5-8 are restricted to the genusGallus but the sequence of one subregion which hybridizes to TBLs is present in the genomes of the order Galliformes.accepted for publication by M. Schmid     

Effects of pentagastrin and carbachol on the gastric histamine level in α-fluoromethylhistidine-treated mice and rats

Summary The gastric mucosal histamine level in mice increased by about 80% and 100% after fasting for 24 and 48 h, respectively. In non-fasted mice, -fluoromethylhistidine (-FMH), a specific histidine decarboxylase inhibitor, significantly decreased the histamine level, the reduction amounting to 35% and 49%, 2 h and 4 h after treatment, respectively. In mice fasted for 24 h, a significant decrease of 42% was observed 4 h after treatment. However, in mice fasted for 48 h, no significant decrease was seen even 4 h after -FMH treatment. Therefore, the histamine-releasing effect of re-feeding and drugs on the gastric mucosa was examined in vivo, using animals fasted for 48 h and subsequently treated with -FMH. Food given simultaneously with -FMH to 48-h fasted mice significantly decreased the histamine level 4 h later. Pentagastrin and carbachol administered alone (0.25–2.0 mg/kg, i.p.) had no significant effect on the histamine level. However, the combined treatment with these drugs significantly decreased the histamine level. In rats fasted for 48 h and treated with -FMH, pentagastrin (0.25 and 0.5 mg/kg, i.p.) but not carbachol (0.125 – 0.5 mg/kg, i. p.) caused a significant decrease in the mucosal histamine level. In contrast to mice, the effect of the combined treatment with pentagastrin and carbachol was not synergistic in rats. These findings suggest that gastrin acts synergistically with acetylcholine in the histamine release from the gastric mucosa in mice, whereas such synergism may not occur in rats. Send offprint requests to K. Saeki     

Prevention of postischemic reperfusion injury: The improvement of myocardial tissue blood flow after ischemia by terminal Nicorandil-Mg cardioplegia

We evaluated the preventive effect of postischemic reperfusion injury by Nicorandil-Mg cardioplegia given just prior to reperfusion as terminal cardioplegia. Twenty seven dogs were placed on cardiopulmonary bypass and the aorta was cross-clamped for 90 min under hypothermic (17–19°C) cardioplegic arrest. The canine hearts were divided into three groups: in group A (n=10) the hearts were reperfused without any treatment; in group B (n=9) the hearts received coronary perfusion with Nicorandil-Mg solution (Nic, 8 mg/l; Mg, 20 mEq/l; glucose, 50 g/l) for 2 min just prior to reperfusion; and in group C (n=8) the hearts received coronary perfusion with Nicorandil-Mg free solution (glucose, 50g/l). During and after ischemia, the myocardial tissue PCO₂ (t-PCO₂) was continuously monitored by an ion-sensitive field effective transistor (ISFET) sensor. In addition, the myocardial tissue blood flow (TBF), oxygen consumption, and lactate flux were then calculated at 5, 10, 20, and 40 min of reperfusion. In the initial reperfusion period, Group B showed an improved TBF compared to group A and C (at 5 min, group B was 42.7±11.9; group A was 29.4±11.2, P<0.025; and group C was 33.9±9.2% of the preischemic control level, P<0.05). T-PCO₂ in group B was significantly decreased at 5 min of reperfusion (group B, 127.5±22.5 42.5±9.7; group A, 117.5±23.0 85.2±17.4, P<0.001; group C, 122.3 mmHg 68.2±18.7 mmHg, P<0.01), and group B had a better metabolic recovery. These results suggest that terminal Nicorandil-Mg cardioplegia might reduce the rate of postischemic reperfusion injury.