Multiple myeloma (MM) is a tumor type characterized by the unregulated proliferation of clonal plasma cells in the bone marrow. Immunotherapy based on chimeric antigen receptor T cell (CAR-T) therapy has achieved exciting success in the treatment of hematological malignant tumors. CD38 is highly and evenly expressed in MM and is an attractive target for MM treatment. Here, we successfully constructed two novel second-generation CAR-T cells targeting CD38 by retroviral vector transduction. CD38 CAR-T cells could be activated effectively after stimulation with CD38-positive tumor cells and secrete cytokines such as IFN-γ and TNF-α to promote tumor cell apoptosis in in vitro experiments. Real-time fluorescence monitoring experiments, luciferase detection experiments and flow cytometry experiments revealed the efficient and specific killing abilities of CD38 CAR-T cells against CD38-positive tumor cells. The proliferation ability of CD38 CAR-T cells in vitro was higher than that of untransduced T cells. Further antitumor experiments in vivo showed that CD38 CAR-T cells could be quickly activated to secrete IFN-γ and eliminate tumors. Thus, novel CD38-targeted second-generation CAR-T cells have efficient and specific antitumor activity and may become a novel therapy for the clinical treatment of MM.

BACKGROUND:cytotoxic lymphocytes are important in the pathogenesis of several disease states, yet, the pathophysiology of lymphocyte-myocyte interaction is not well known.METHODS AND RESULTS:We have developed a model for the in vitro evaluation of autoimmune cytotoxic myocardial damage. Cardiac myocytes were repeatedly injected to adult autologous rats. Following 3 months, histological evidence of myocarditis was seen in 20% of the hearts. Cultured myocytes obtained from newborn rats were exposed to lymphocytes isolated from the immunized animals. Cytotoxic activity was measured using crystal violet staining test. The percentage of killing was increased as the ratio of lymphocytes/myocytes was increased. Verapamil did not block this cytotoxic effect. No killing was seen when myocytes were exposed to non-sensitized lymphocytes. Physiological changes induced in myocytes by cytotoxic lymphocytes were studied. Cell wall motion was measured by an optical method and action potentials with intracellular microelectrodes. Physiological changes observed in myocytes following exposure to cytotoxic lymphocytes included: Impaired relaxation with prolonged contractions, oscillations and prolongation of the plateau of the action potential. Cellular contraction was prolonged up to 4 s before total arrest of spontaneous activity. Verapamil but not tetrodotoxin restored action potentials and contractions to normal. Supernatant collected from cultures of myocytes and lymphocytes had the same effect on myocytes contractility as observed following exposure of myocytes to cytotoxic lymphocytes.CONCLUSIONS:This supports our hypothesis that these physiological alterations observed in myocytes are mediated by a soluble factor secreted by cytotoxic lymphocytes.

Patients with diabetes mellitus are excessively vulnerable to myocardial ischemia and often suffer from autonomic cardiac dysfunction. They are also known to have specific capillary pathology and abnormalities of substances essential for the neoangiogenic cascade. These background data led us to the hypothesis that the severity of ischemic heart disease in diabetes is attributable (at least in part) to microcirculatory and autonomic inadequacy caused by microangiopathy and failure of postischemic adaptive neoangiogenesis. To test this hypothesis we compared myocardial capillaries, autonomic nerve endings of 19 diabetics, 30 normoglycemics with ischemic heart disease, and 9 valve replacements serving as nonischemic controls. Right atrial appendages obtained during coronary bypass surgery were utilized for light, fluorescent, and electron microscopic morphometry. Although in this series there were no significant differences in the clinical and laboratory hemodynamic values between the ischemic normoglycemic and diabetic patients, the latter showed marked capillary and nerve terminal pathology, and their capillary density as well as capillary to myofiber ratios were significantly lower. In addition, the mean capillary area, volume fraction, and intercapillary distance were higher in diabetics. The mean area of the nerve varicosities was also smaller, and this was correlated with capillary density. We concluded that the findings support the hypothesis that cardiac vulnerability in diabetes is connected with inadequate adaptive neoangiogenesis and that this seems to be associated with atrophic changes in the nerve terminals.

Down's Syndrome (DS), the phenotypic expression of human trisomy 21, is presumed to result from overexpression of certain genes residing on chromosome 21 at the segment 21q22-the Down locus. The "housekeeping" enzyme CuZn-superoxide dismutase (CuZnSOD) is encoded by a gene from that region and its activity is elevated in DS patients. Moreover, the recent discovery that familial ALS is associated with mutations in the gene encoding CuZnSOD, focused attention on the entanglement of oxygen-free radicals in cell death and neuronal disorders. To investigate the involvement of CuZnSOD gene dosage in the etiology of the syndrome we have developed both cellular and animal models which enabled us to investigate the physiological consequences resulting from overexpression of the CuZnSOD gene. Rat PC12 cells expressing elevated levels of transfected human CuZnSOD gene were generated. These transformants (designated PC12-hSOD) closely resembled the parental cells in their morphology, growth rate, and response to nerve growth factor, but showed impaired neurotransmitter uptake. The lesion was localized to the chromaffin granule transport mechanism. These results show that elevation of CuZnSOD activity interferes with the transport of biogenic amines into chromaffin granules. Since neurotransmitter uptake plays an important role in many processes of the central nervous system, CuZnSOD gene-dosage may contribute to the neurobiological abnormalities of Down's Syndrome. As an approach to the development of an animal model for Down's Syndrome, several strains of transgenic mice which carry the human CuZnSOD gene have been prepared. These animals express the transgene as an active enzyme with increased activity from 1.6 to 6.0-fold in the brains of four transgenic strains and to an equal or lesser extent in several other tissues. To investigate the contribution of CuZnSOD gene dosage in the neuropathological symptoms of Down's Syndrome, we analyzed the tongue muscle of the transgenic-CuZnSOD mice. The tongue neuromuscular junctions (NMJ) in the transgenic animals exhibited significant pathological changes; withdrawal and destruction of some terminal axons and the development of multiple small terminals. The ratio of terminal axon area to postsynaptic membranes decreased, and secondary folds were often complex and hyperplastic. The morphological changes in the transgenic NMJ were similar to those previously seen in the transgenic NMJ and were similar to those previously seen in muscles of aging mice and rats as well as in tongue muscles of patients with Down's Syndrome. The findings suggest that CuZnSOD gene dosage is involved in the pathological abnormalities of tongue NMJ observed in Down's Syndrome patients.(ABSTRACT TRUNCATED AT 400 WORDS)

1. Light and electron microscope morphometry has been used to study the effect on heart morphology of a sedentary life style in normothermic (24-degrees-C) and warm acclimated (34-degrees-C) rats in comparison with exercised normothermic and warm acclimated rats.2. The sedentary acclimated rats had multiple foci of myocardial necrosis while the exercised rats did not. The capillary to myofibre ratio and the number of mast cells were higher in the acclimated exercised hearts than in hearts from all the other groups. The intercapillary distance and the mitochondrial volume fractions were not affected by any treatment regime.3. It is concluded that although some neoangiogenesis occurs during exercise + heat, the neoangiogenesis does not explain the absence of myocardial pathology produced by heat alone.

A375 human melanoma cell cultures grown in the presence of TGF-beta contained greatly reduced cell numbers and exhibited drastic alterations in cell morphology compared to the control cultures. Preincubation of the cells with the cytokine for only 18 h was sufficient to induce these changes irreversibly. Examination of TGF-beta-treated cells in the electron microscope revealed large numbers of lipid-filled vacuoles in the cytoplasm, greatly contracted nuclei and some loss of the otherwise abundant microvilli. Thus TGF-beta may have a direct toxic effect on the A375 melanoma cells.

BACKGROUND:The coronary microvasculature may be abnormal even in the presence of angiographically normal epicardial arteries. Abnormalities of small coronary vessels have been invoked as a cause of angina.METHODS AND RESULTS:To quantitatively evaluate the morphology of capillaries in patients with idiopathic dilated cardiomyopathy (DCM) or the syndrome of angina and small vessel disease (SVD), we performed electron microscopic morphometry of capillaries in right ventricular biopsy samples taken from 32 patients. Ten had angina, normal epicardial coronary arteries, and evidence for SVD; 12 had DCM; and 10 had normal hearts. In patients with DCM, the ratio of microvessels to myocytes was not different than that of controls (0.49 +/- 0.06 versus 0.51 +/- 0.05). Mean cross-sectional areas of the capillaries (lumen plus wall) and lumen were significantly greater than those of controls (45.3 +/- 15.1 versus 22.7 +/- 8.3 micron 2, p less than 0.001; 17.6 +/- 6.9 versus 11.6 +/- 6.2 micron 2, p less than 0.05, respectively). Fibrous content of the myocardium, as assessed by quantitative light microscopy, was significantly increased (16.3 +/- 3.3% versus 5.0 +/- 2.4%, p less than 0.001). In contrast, in patients with SVD, the capillary-to-myocyte ratio was reduced (0.33 +/- 0.08, p less than 0.001). Although mean cross-sectional areas of the entire capillary (32.4 +/- 19.7 micron 2) and the lumen (8.9 +/- 7.8 micron 2) were not statistically different than those of controls, there was an absence of capillaries less than 15 micron 2 in cross-sectional area, and the frequency distribution of the lumen area was skewed to the left. Swollen endothelial cells frequently encroached upon the lumen. There was a mild increase in fibrous content (9.5 +/- 3.7%, p less than 0.05).CONCLUSIONS:Enlarged capillaries and a normal ratio of capillaries to myocytes appear to be features of DCM. Of the patients with SVD, there was both a relative lack of capillaries and capillary lumen narrowing from swollen endothelium. These changes may induce ischemia and angina and may result in mild fibrosis.

Radiation damage to the neuromuscular junctions (NMJs) in mouse tongues was studied using local x-irradiation of the tongues with the rest of the body shielded. Transmission electron microscopy (TEM) revealed no significant morphological changes in the fine structures and organelles of the NMJs given 4 Gy. A dose of 8 Gy produced degenevative morphological changes associated with oxon terminal sprouting as early as 2 and 7 days following irradiation. Subsequently, 1-11 weeks later, severe degenerative changes were observed. The number of mitochondria was significantly decreased with increased occurrence of degenerative membranal features. The number of synaptic footplates without terminals or with multiple small terminals within one groove increased gradually with time. Most of these pathological changes persisted for at least 3 months after irradiation. However, the myofibres, blood vessels and interstitial cells appeared to be unaffected throughout the period of follow-up. The present study substantiates our previous reports of ageing-like changes in the tongues' NMJs induced by their excessive exposure to free radicals.

This study focuses on the ultrastructure of microvessels and myofibers in 54 endomyocardial biopsy specimens from 16 patients treated with cyclosporine after heart transplantation. Although the capillary and myofiber pathologic condition was significantly correlated with the degree of rejection, ultrastructural changes sometimes occurred in biopsy specimens with little or no histologic cellular infiltration. Immunocytochemical studies of some biopsy specimens showed that all microvessels were positive for IgM, whereas concentrations of IgG, complement, and fibrin varied. The microvascular and myofiber changes appeared to be reversible and correlated (Pearson's coefficient of correlation of myofiber vacuoles and microangiopathy, R = 0.67, p less than 0.00001). The vacuoles in myofibers resembled those in subendocardial regions of ischemic myocardium. They could, however, be related to cyclosporine toxicity because similar vacuoles have been described in kidneys of transplant patients receiving cyclosporine. We conclude that in cases of unclear graft failure in the absence of classic signs of rejection, electron microscopy could be helpful in detecting microvascular and myofiber pathologic situations that may influence myocardial function.

Down syndrome (DS), the phenotypic expression of human trisomy 21, is presumed to result from overexpression of certain genes residing on chromosome 21 at the segment 21q22-the Down locus. The "housekeeping" enzyme CuZn-superoxide dismutase (CuZnSOD) is encoded by a gene from that region and its activity is elevated in DS patients. To investigate the possible involvement of CuZnSOD gene dosage in the etiology of the syndrome we have developed both cellular and animal models which enabled us to investigate the physiological consequences resulting from overexpression of the CuZnSOD gene. 1. Rat PC12 cells expressing elevated levels of transfected human CuZnSOD gene were generated. These transformants (designated PC12-hSOD) closely resembled the parental cells in their morphology, growth rate, and response to nerve growth factor, but showed impaired neurotransmitter uptake. The lesion was localized to the chromaffin granule transport mechanism. We found that the pH gradient (delta pH) across the membrane, which is the main driving force for amine transport, was diminished in PC12-hSOD granules. These results show that elevation of CuZnSOD activity interferes with the transport of biogenic amines into chromaffin granules. Since neurotransmitter uptake plays an important role in many processes of the central nervous system, CuZnSOD gene-dosage may contribute to the neurobiological abnormalities of Down's syndrome. 2. As an approach to the development of an animal model for Down syndrome, several strains of transgenic mice that carry the human CuZnSOD gene have been prepared. These animals express the transgene in a manner similar to that of humans, with 0.9 and 0.7-kilobase transcripts in a 1:4 ratio, and synthesize the human enzyme in an active form capable of forming human-mouse enzyme heterodimers. CuZnSOD activity is increased from 1.6 to 6.0-fold in the brains of four transgenic strains and to an equal or lesser extent in several other tissues. 3. To investigate the possible involvement of CuZnSOD gene dosage in the neuropathological symptoms of Down's syndrome, we analyzed the tongue muscle of the transgenic mice that express elevated levels of human CuZnSOD. The tongue neuromuscular junctions (NMJ) in the transgenic animals exhibited significant pathological changes, namely, withdrawal and destruction of some terminal axons and the development of multiple small terminals. The ratio of terminal axon area to postsynaptic membrane decreased, and secondary folds were often complex and hyperplastic. The morphological changes in the transgenic NMJ were similar to those previously seen in muscles of aging mice and rats as well as in tongue muscle of patients with Down's syndrome.(ABSTRACT TRUNCATED AT 400 WORDS)

6 patients, 37-55 years old, who underwent heart transplantation in Belgium, have been followed in our clinic for the past 2 years. All had suffered for 6-12 months from congestive heart failure which had not responded to intensive medical treatment and were considered terminal. 5 had had ischemic heart disease and 1 idiopathic dilated cardiomyopathy. After transplantation all patients were again in good physical condition and 5 of them had resumed a productive lifestyle. In 3, follow-up endomyocardial biopsies revealed episodes of "moderate rejection" of the transplant, all of which were successfully treated with pulse courses of corticosteroids and increased cyclosporine dosage. Mild renal failure developed in 3, and in 3 systemic hypertension, controlled by antihypertensive drugs. 4 patients examined in our department and found suitable for transplantation died before a compatible donor was found. In 2 other patients heart transplantation was not possible due to severe pulmonary hypertension. Heart transplantation is recommended for patients with terminal congestive heart failure unresponsive to medical treatment. Early transplantation prevents irreversible changes in the pulmonary vessels which would make operation impossible, and lowers operative risk.

To investigate the possible involvement of Cu/Zn-superoxide dismutase (CuZnSOD) gene dosage in the neuropathological symptoms of Down's syndrome, we analyzed the tongue muscle of transgenic mice that express elevated levels of human CuZnSOD. The tongue neuromuscular junctions (NMJ) in the transgenic animals exhibited significant pathological changes, namely, withdrawal and destruction of some terminal axons and the development of multiple small terminals. The ratio of terminal axon area to postsynaptic membrane decreased, and secondary folds were often complex and hyperplastic. The morphological changes in the transgenic NMJ were similar to those previously seen in muscles of aging mice and rats as well as in tongue muscle of patients with Down's syndrome. The findings suggest that CuZnSOD gene dosage is involved in the pathological abnormalities of tongue NMJ observed in Down's syndrome patients.

Cationized ferritin binding was used to measure negative surface charge on endothelium of large arteries in streptozotocin-induced diabetic rats and normal control rats. The negative charge was significantly lower in the diabetic animals (p less than 0.01). This change, possibly related to glycosylation, may lead to altered vascular permeability and may be of importance in the vascular pathology of diabetes.

We examined the tongue muscles in several strains of transgenic mice carrying the human Zn-Cu superoxide dismutase (CuZnSOD) gene. The presence of the extra gene was confirmed in mated progeny and the gene product activity was measured in the tongue and found to be much higher than in normal littermate controls. Using electron microscopic morphometry, the neuromuscular junctions of the transgenic mice showed significant changes resembling excessive aging, with atrophy, degeneration, withdrawal, and sometimes destruction of the terminal axons, as well as the development of multiple small terminals. The myofibers showed little change except for slight hypertrophy and an increased variability in size. They also had more megamitochondria, fat droplets and lipofuscin bodies. Excess CuZnSOD generates H2O2 and hydroxyl radicals which affect both NMJ membranes and plasticity, and which may produce premature aging. The findings resemble those observed in tongues of patients with Down's syndrome, in whom an extra CuZnSOD gene is present as part of the trisomy of chromosome 21.