An electron microscopic and immunohistochemical study of Merkel cell carcinoma

Two cases of Merkel cell carcinoma (MCC) were examined by electron microscopy and immunohistochemistry. Histologically, tumor cells, which extended from the dermis into the subcutis, showed anastomosing bands with partial trabecular pattern. The ultrastructural study showed tumor cells in case 1 with numerous neurosecretory granules. The number of granules in case 2, however, was smaller compared with that in case 1. Perinuclear bundles of filaments were present in case 2, but few bundles were observed in case 1. By immunohistochemistry, cytokeratin (CK)-8, -18, -19, and -20 and epithelial membrane antigen were stained positively within tumor cells in both cases. It was interesting that staining patterns of chromogranin A and of neuron-specific enolase were different in the two cases. These data indicated that CK-20 is a useful marker for diagnosing MCC and that ultrastructural and immunohistochemical differences in both cases were the result of phenotypic variation.     

Cytotoxic antimelanoma drugs suppress the activation of M2 macrophages

Together with regulatory T cells (Tregs), tumor‐associated macrophages (TAMs) play roles in maintaining the tumor microenvironment. Although cytotoxic antimelanoma drugs such as dacarbazine (DTIC), nimustine hydrochloride (ACNU) and vincristine (VCR) have been used for the treatment of malignant melanoma as adjuvant therapy in Japan, the detailed mechanisms of their immunomodulatory effects are not fully understood. As the majority of TAMs are alternatively activated M2 macrophages that favour tumor development, the aim of this study was to elucidate the immunomodulatory effects of these reagents on human monocyte‐derived M2 macrophages. First, mRNA expressions and protein production of immune checkpoint molecules, PD‐L1 and chemokines by CD163⁺ CD206⁺ M2 macrophages derived from peripheral blood mononuclear cells were investigated to determine the immunomodulatory effects of DTIC, ACNU, and VCR. DTIC and VCR significantly decreased PD‐L1 mRNA expression, which was confirmed by flow cytometry. Moreover, the mRNA expression and production of CCL22 were significantly decreased by DTIC, which suggested that DTIC might suppress the recruitment of Tregs in the tumor site. Furthermore, the decreased expression of PD‐L1 and production of CCL22 were validated in vivo, using the B16F10 mouse melanoma model, leading to abrogation of the suppressive function of T‐cell proliferation. The present report suggests one of the possible antimelanoma mechanisms of DAV combination chemotherapy for melanoma patients.     

Spin labeling of a cysteine residue of the Escherichia coli outer membrane lipoprotein in its membrane environment

A method was developed to attach a spin label to a specific site on the structural lipoprotein of the Escherichia coli outer membrane in situ. This method takes advantage of the fact that the outer membrane of wild-type E. coli contains few residues reactive towards sulfhydryl reagents. A mutant E. coli strain has been isolated [Suzuki, H., Nishimura, Y., Iketani, H., Campisi, J., Hirashima, A., Inouye, M. & Hirota, Y. (1976) J. Bacteriol. 127, 1494-1501] in which the second position from the carboxy terminus of the lipoprotein is changed from arginine into a cysteine residue. The membrane fraction of this mutant was treated with N-(1-oxyl-2,2,5,5-tetramethylpyrrolidinyl)maleimide in the presence of EDTA and 2-mercaptoethanol. Spin label was found to be preferentially incorporated into the lipoprotein. The spectrum of the spin-labeled membrane shows two components, both arising from spin label at the same site near the carboxy terminus. The strongly immobilized component has a maximum hyperfine splitting value of 53 G, and the weakly immobilized component, 37 G. A fraction of the lipoprotein is covalently bound to the peptidoglycan layer through its carboxy-terminal lysine; the spectrum of the isolated bound form of the lipoprotein was identical to that of the free form. When the matrix protein, the other major outer membrane protein, was removed by mutation, the spectrum of the lipoprotein was altered, suggesting that these two proteins are closely associated.     

Altered Cerebellum Development and Dopamine Distribution in a Rat Genetic Model with Congenital Hypothyroidism

Thyroid hormones play crucial roles in the development and functional maintenance of the central nervous system. Despite extensive studies of the neural function of thyroid hormones, little is known about the effects of hypothyroidism on behavioural traits and the mechanisms underlying such effects. In the present study, we report an investigation of congenitally hypothyroid mutant rdw rats, revealing a novel function of thyroid hormones in the central nervous system. The rdw rats were subjected to behavioural analyses such as the rotarod test, open field test and circadian activity measurement. To determine the cause of behavioural disorders, cerebellar morphogenesis was examined by immunohistochemical analysis, and the axonal transport of dopamine in the nigrostriatal pathway was analysed by high‐performance liquid chromatography and western blotting. The effects of thyroxine administration to the rdw rats were examined by behavioural analysis. The rdw rats showed severe impairment of motor coordination and balance. This could be explained by the fact that the rats showed severe retardation of cerebellar morphogenesis, which correlates with the small somata and poor dendritic arborisation of Purkinje cells and retarded migration of granule cells particularly during the first two postnatal weeks. Moreover, the rdw rats showed hypoactivity, characterised by decreased circadian locomotor activity. After weaning, thyroxine administration improved the dwarfism in rdw rats but had no effect on cerebellar function. In addition, the rdw rats showed anxiety and depression intrinsically to novel surroundings. Interestingly, the rdw rats showed high levels of dopamine in the substantia nigra and low levels in the striatum, an important centre for the coordination of behaviour. Furthermore, low levels of tubulin in the striatum were detected, indicating the aberrant axonal transport of dopamine in the nigrostriatal pathway as a result of the reduced delivery of microtubules. These findings indicate an important function of thyroid hormones in cerebellar formation and in the regulation of axonal transport of dopamine. Moreover, rdw rats will be useful for studies of brain function and behavioural disorders in congenital hypothyroidism.     

Real-time tumor-tracking radiotherapy for adrenal tumors.

PURPOSE: To investigate the three-dimensional movement of internal fiducial markers near the adrenal tumors using a real-time tumor-tracking radiotherapy (RTRT) system and to examine the feasibility of high-dose hypofractionated radiotherapy for the adrenal tumors. MATERIALS AND METHODS: The subjects considered in this study were 10 markers of the 9 patients treated with RTRT. A total of 72 days in the prone position and 61 treatment days in the supine position for nine of the 10 markers were analyzed. All but one patient were prescribed 48 Gy in eight fractions at the isocenter. RESULTS: The average absolute amplitude of the marker movement in the prone position was 6.1+/-4.4 mm (range 2.3-14.4), 11.1+/-7.1 mm (3.5-25.2), and 7.0+/-3.5 mm (3.9-12.5) in the left-right (LR), craniocaudal (CC), and anterior-posterior (AP) directions, respectively. The average absolute amplitude in the supine position was 3.4+/-2.9 mm (0.6-9.1), 9.9+/-9.8 mm (1.1-27.1), and 5.4+/-5.2 mm (1.7-26.6) in the LR, CC, and AP directions, respectively. Of the eight markers, which were examined in both the prone and supine positions, there was no significant difference in the average absolute amplitude between the two positions. No symptomatic adverse effects were observed within the median follow-up period of 16 months (range 5-21 months). The actuarial freedom-from-local-progression rate was 100% at 12 months. CONCLUSIONS: Three-dimensional motion of a fiducial marker near the adrenal tumors was detected. Hypofractionated RTRT for adrenal tumors was feasible for patients with metastatic tumors.     

Dosimetry of fission neutrons in a 1-W reactor,UTR-KINKI

The energy spectrum of fission neutrons in the biological irradiation field of the Kinki University reactor, UTR-KINKI, has been determined by a multi-foil activation analysis coupled with artificial neural network techniques and a Au-foil activation method. The mean neutron energy was estimated to be 1.26 +/- 0.05 MeV from the experimentally determined spectrum. Based on this energy value and other information, the neutron dose rate was estimated to be 19.7 +/- 1.4 cGy/hr. Since this dose rate agrees with that measured by a pair of ionizing chambers (21.4 cGy/hr), we conclude that the mean neutron energy could be estimated with reasonable accuracy in the irradiation field of UTR-KINKI.     

Prognostic value of impaired myocardial fatty acid uptake in patients with acute myocardial infarction

Impaired cardiac fatty acid uptake, assessed by the radiolabelled fatty acid analogue beta-methyl-p-iodophenyl pentadecanoic acid (I-123-BMIPP), is observed in the myocardium following acute ischaemic events, but the long-term prognostic implication has not been established. This study aimed to determine the prognostic value of cardiac BMIPP uptake in patients with acute myocardial infarction. Following the assessment of thallium-201 and I-123-BMIPP uptake, 101 post-infarct patients were prospectively followed up with primary end points of cardiac death, heart failure and non-fatal infarction. During a mean follow-up of 28 months, three cardiac deaths, three non-fatal infarctions, 23 revascularizations and four recurrences of angina pectoris were observed. Multivariate analysis identified reduced uptake of BMIPP and perfusion, no beta-blocking treatment and greater thallium-BMIPP mismatch (i.e. larger BMIPP defect than thallium defect) as significant predictors for overall cardiac events. Prior myocardial infarction, reduced left ventricular ejection fraction and greater thallium-BMIPP mismatch were selected as independent predictors of harder cardiac events. Female patients, those with greater BMIPP defect or greater thallium-BMIPP mismatch showed worse clinical outcomes. The inclusion of BMIPP data improved the prognostic values of conventional significant predictors. In conclusion, impaired myocardial I-123-BMIPP uptake and perfusion-BMIPP mismatch are related to a high probability of fatal and non-fatal cardiac events, suggesting the aetiological relevance and prognostic value of impaired cardiac fatty acid metabolism in viable, but jeopardized, myocardium following acute myocardial infarction.