Effects of surface charge and palladium on hepatic and kidney injury induced by polystyrene nanoparticles co-administered to mice with paraquat and cisplatin.

Recently, with the advancement of nanotechnology, various nanoparticles have been developed and used in fields such as electronics, cosmetics, and foods. However, the toxicity of nanoparticles has yet to be fully investigated. In particular, the interactions between nanoparticles and therapeutic drugs require further study. We previously reported that unmodified polystyrene nanoparticles with a particle size of 50 nm (NPP50) co-administered with paraquat (PQ) or cisplatin (CDDP) induce hepatic and kidney injury. Here, we determined if NPP50 modified with the amino group (NPP50-NH2), carboxyl group (NPP50-COOH), or palladium (Pd-NPP50) caused liver or kidney injury when co-administered with PQ or CDDP. The results showed that when NPP50-NH2, NPP50-COOH, or Pd-NPP50 was administered alone via the mouse tail vein, serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and blood urea nitrogen (BUN) did not increase or cause injury. When NPP50, NPP50-NH2, NPP50-COOH, or Pd-NPP50 was co-administered with PQ, serum levels of ALT and AST increased in the NPP50 group but did not increase in the NPP50-NH2, NPP50-COOH, or Pd-NPP50 groups. When NPP50-NH2, NPP50-COOH, or Pd-NPP50 was co-administered with CDDP, ALT, AST, and BUN values did not increase. These data suggest that injury due to the interaction of polystyrene nanoparticles with CDDP or PQ can be suppressed by changes in the surface charge of nanoparticles or by Pd modification.

The hepatoprotective effect of lycopene on Con A-induced liver injury in mice.

Lycopene, the main fat-soluble pigment responsible for the red color of ripe tomatoes, is a symmetrical tetraterpene comprising eight isoprene units. In vitro and in vivo studies have shown that lycopene acts as a potent antioxidant; it is 100 times more effective than vitamin E and 125 times more effective than glutathione as an antioxidant. Here, we divided BALB/c male mice into three equal groups: control, Concanavalin A (Con A), and Con A and lycopene. The control group mice received only vehicle by intraperitoneal injection, the Con A group mice were given Con A, and the Con A and lycopene group mice received Con A and lycopene. The results showed that Con A administration increased histopathological damage, and the levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), interleukin (IL)-6, interferon (IFN)-γ and tumor necrosis factor (TNF)-α were increased in serum samples whereas the levels of these compounds were significantly decreased in the Con A and lycopene group compared to the Con A group. Furthermore, we observed that lycopene led to an increase in cell viability and cell growth. The results of this study revealed that lycopene might be a useful hepatoprotective agent for reducing increased proinflammatory cytokine levels, and for increasing cell viability and cell growth.

Hepatotoxicity, nephrotoxicity, and drug/chemical interaction toxicity of platinum nanoparticles in mice.

Nanomaterials are frequently used in microelectronics, cosmetics, and sunscreens. Platinum reagents are commonly used in disease diagnosis, cosmetics, and the food industry. Although research into the development of nanomaterialbased drug delivery systems has yielded promising results, the toxicity of these materials is not fully understood. We investigated the toxicity and drug interactions of 1- and 8-nm diameter platinum nanoparticles (nPt1 and nPt8, respectively) in mice. Acute hepato-renal toxicity of intravenously administered platinum nanoparticles was evaluated biochemically and histologically. Dose-dependent increases in serum markers of hepato-renal function (serum aminotransferases and blood urea nitrogen) were observed following administration of nPt1, whereas nPt8 had no effect, even at 20 mg/kg. Moreover, nPt1 induced interleukin (IL)-6 and IL-1ß production 3 and 6 hours after administration. The effect of nPts on drug-induced toxicity was evaluated in mice injected intraperitoneally with carbon tetrachloride or cisplatin, with or without intravenous administration of platinum nanoparticles. All treatments in the absence of nanoparticles were non-lethal and resulted in moderate toxicity. However, exacerbated toxicity was observed in mice injected with carbon tetrachloride or cisplatin together with nPt1, but not in mice co-injected with nPt8. We found that nPt1 cause hepato-renal damage, and the effect is enhanced by chemical inducers of hepatotoxicity and nephrotoxicity. This is the first report demonstrating that nPt1 not only are hepatotoxic and nephrotoxic but also exacerbate drug toxicity. These findings will be useful for future nanotechnology and nanoscience research.

Toxicity of 50-nm polystyrene particles co-administered to mice with acetaminophen, 5-aminosalicylic acid or tetracycline.

We investigated whether nano-sized polystyrene particles affect drug-induced toxicity. The particles, which are widely used industrially, had diameters of 50 (NPP50), 200 (NPP200) or 1000 (NPP1000) nm. The toxic chemicals tested were acetaminophen (APAP), 5-aminosalicylic acid (5-ASA), tetracycline (TC), and sodium valproate (VPA). All treatments in the absence of the nanoparticles were non-lethal and did not result in severe toxicity. However, when mice were injected with APAP, 5-ASA or TC together with polystyrene particles, synergistic, enhanced toxicity was observed in mice injected with NPP50. These synergic effects were not observed in mice co-injected with NPP200 or NPP1000. On the other hand, co-administration of VPA and NPP50, NPP200 or NPP1000 did not elevate toxicity. The results show that NPP50 differs in hepatotoxicity depending on the drug co-administered. These findings suggest that further evaluation of the interactions between polystyrene nanoparticles and drugs is a critical prerequisite to the pharmaceutical application of nanotechnology.

Functional expression and germline transmission of a human chromosome fragment in chimaeric mice.

Human chromosomes or chromosome fragments derived from normal fibroblasts were introduced into mouse embryonic stem (ES) cells via microcell-mediated chromosome transfer (MMCT) and viable chimaeric mice were produced from them. Transferred chromosomes were stably retained, and human genes, including immunoglobulin (Ig) kappa, heavy, lambda genes, were expressed in proper tissue-specific manner in adult chimaeric tissues. In the case of a human chromosome (hChr.) 2-derived fragment, it was found to be transmitted to the offspring through the germline. Our study demonstrates that MMCT allows for introduction of very large amounts of foreign genetic material into mice. This novel procedure will facilitate the functional analyses of human genomes in vivo.

Influence of 50-nm polystyrene particles in inducing cytotoxicity in mice co-injected with carbon tetrachloride, cisplatin, or paraquat.

The toxicity of nanomaterials has yet to be fully investigated. In particular, the interactions between nanomaterials and therapeutic drugs require further study. We investigated whether nano-sized polystyrene particles affect drug-induced toxicity. The particles, which are widely used industrially, had diameters of 50 (NPP50), 200 (NPP200) or 1000 (NPP1000) nm. The toxic chemicals tested were carbon tetrachloride, cisplatin (a popular anti-tumor agent), and a widely used herbicide, paraquat. Mice were treated intraperitoneally with either carbon tetrachloride (0.01 ml/kg), cisplatin (100 micromol/kg) or paraquat (50 mg/kg), with or without intravenous administration of polystyrene particles. All treatments in the absence of the nanoparticles were non-lethal and did not result in severe toxicity. However, when mice were injected with paraquat or cisplatin together with polystyrene particles, synergistic, enhanced toxicity was observed in mice injected with NPP50. These synergic effects were not observed in mice co-injected with NPP200 or NPP1000. These findings suggest that further evaluation of the interactions between polystyrene nano-particles and drugs is a critical prerequisite to the pharmaceutical application of nanotechnology.

Characterization of human-type monoclonal antibodies against reduced form of hemin binding protein 35 from Porphyromonas gingivalis.

BACKGROUND AND OBJECTIVE: The gram-negative anaerobe Porphyromonas gingivalis has been implicated as an important pathogen in the development of adult periodontitis, and its colonization of subgingival sites is critical in the pathogenic process. We previously identified a 35 kDa surface protein (hemin binding protein 35; HBP35) from P. gingivalis that exhibited coaggregation activity, while additional analysis suggested that this protein possessed an ability to bind heme molecules. For development of passive immunotherapy for periodontal diseases, human-type monoclonal antibodies have been prepared using HBP35 as an antigen in TransChromo mice. In the present study, we focused on a single antibody, TCmAb-h13, which is known to inhibit heme binding to recombinant HBP35. The aim of our investigation was to clarify the redox-related function of HBP35 and consider the benefits of human-type monoclonal antibodies. MATERIAL AND METHODS: To examine the antigen recognition capability of TCmAbs with immunoblotting and Biacore techniques, we used the native form as well as several Cys-to-Ser variants of recombinant HBP35. RESULTS: We found that the redox state of recombinant HBP35 was dependent on two Cys residues, (48) C and (51) C, in the thioredoxin active center (WCGxCx). Furthermore, TCmAb-h13 recognized the reduced forms of recombinant HBP35, indicating its inhibitory effect on P. gingivalis growth. CONCLUSION: Hemin binding protein 35 appears to be an important molecule involved in recognition of the redox state of environmental conditions. In addition, TCmAb-h13 had an inhibitory effect on heme binding to recombinant HBP35, thereby interfering with P. gingivalis growth.

Transgenic mice demonstrate that epithelial homing of gamma/delta T cells is determined by cell lineages independent of T cell receptor specificity.

gamma/delta T cells with different TCR repertoires are compartmentalized in different epithelia. This raises the possibility that the TCR-gamma/delta directs homing of T cells to these epithelia. Alternatively, the signals that induce TCR-gamma/delta expression in developing T cells may also induce homing properties in such cells, presumably in the form of cell surface receptors. We have examined this issue by studying the homing of gamma/delta T cells in transgenic mice constructed with specific pairs of rearranged gamma and delta genes. In such mice, most gamma/delta T cells express the transgene-encoded TCR. We find that homing to both skin and gut epithelia is a property of T cells and is not determined by the type of gamma and delta genes used to encode their TCR. We also studied the effect of TCR replacement on the expression of Thy-1 and CD8 proteins on the gamma/delta T cells associated with gut epithelia. Our results show that the expression of the appropriate type of TCR-gamma/delta is not required for the Thy-1 expression by these T cells, suggesting that Thy-1 is not an activation marker. In contrast, CD8 expression by gut gamma/delta T cells seems to depend on the expression of the appropriate type of TCR.

[Radical surgery using modified open door method for anterior apical tumor].

The approach that should be used for an anterior apical tumor still remains controversial. Since a modified open door method was very useful for the widening of the surgical field in a recent patient with an anterior apical tumor, an outline of this case is reported. The patient was a 66-year-old male with squamous cell carcinoma of the anterior apical region of the right lung (suspected to be invading the thoracic wall, cT3N1M0). After a midline sternal incision with a right unilateral collar incision, the medial half of the right clavicle and a few cm of the right 1st rib on the sternal side were resected to sufficiently expose the area from the right brachiocephalic trunk to around the subclavicular artery and vein, where invasion was suspected. This treatment facilitated widening of the visual field around the site of tumor invasion and made safe right upper lobectomy + combined thoracic wall resection + ND2a possible. In this patient, anterolateral incision at the 4th intercostal level, which is made using the original open door method, could be avoided, probably minimizing surgical invasion.

Human monoclonal antibody for ovarian clear cell carcinoma-2, a human monoclonal antibody with antitumor activity against ovarian cancer cells that recognizes CA125-like antigen.

In recent years, antibody therapy employing monoclonal antibodies has become a new approach for treating cancer. This study was performed to establish a human monoclonal antibody recognizing an epitope related to CA125 using KM mice and to assess its reactivity with ovarian cancer cells. A human ovarian clear cell adenocarcinoma cell line (RMG-I) was used to immunize KM mice, and hybridoma supernatant was obtained by a standard method employing enzyme-linked immunosorbent assay screening. Next, selection of hybridomas was performed with two antibodies (MA602-1 and MA602-6) and a sandwich immunoassay for CA125-like antigen, and then the limiting dilution was used to obtain a human monoclonal antibody. Immunohistochemical reactivity of this antibody (human monoclonal antibody for ovarian clear cell carcinoma-2 [HMOCC-2]) with ovarian cancer was assessed, while its specificity was analyzed by Western blotting. Various antibodies were used to identify the epitope targeted by HMOCC-2. Finally, the antitumor effect of HMOCC-2 was assessed by intraperitoneal administration to SCID (severe combined immunodeficiency) mice with heterografts of RMG-I tumors. HMOCC-2 showed a positive reaction with 60% (63/105) of ovarian cancer specimens. Western blotting of the membrane fraction of RMG-I revealed several bands at 120-250 kd. HMOCC-2 recognized the CA125-like antigens identified by several antibodies. HMOCC-2 also exhibited significant antitumor activity (P < 0.01) against ovarian cancer heterografts. HMOCC-2 reacts specifically with ovarian cancer cells via a target epitope analogous to that of CA125 and also exhibits activity against ovarian tumors. These findings suggest that it may have the potential to be employed clinically for molecular-targeting therapy.

[Sternal resection and chest wall reconstruction for primitive neuroectodermal tumor of the sternum].

Primitive neuroectodermal tumor of the sternum is rare. A 59-year-old woman referred to our department with anterior chest pain and a tumor in the sternum. The patient was diagnosed as primitive neuroectodermal tumor of the sternum by core biopsy of the lesion. She received 2 cycles of preoperative chemotherapy with vincristine, doxorubicin, cyclophosphamide, ifosfamide, etoposide. She underwent a total sternectomy with resection of adjacent bilateral costal cartilages and sternal ends of the clavicles. The skeletal defect of chest wall was reconstructed by polypropylene mesh-resin sandwich. The myocutaneus defect was reconstructed by the pedicled latissimus dorsi myocutaneus flap and the bilateral breast flaps. The postoperative course was uneventful and adjuvant radiotherapy was started 6 weeks after the operation. She died of distant metastases 3 months after the operation, although this patient was free from local recurrence.

Virus-induced cell death in plants expressing the mammalian 2',5' oligoadenylate system.

The major components of the 2-5A system, responsible for the mammalian interferon-induced antiviral response, are the 2',5' oligoadenylate synthetase (2-5Aase) and 2',5' oligoadenylate (2-5A) dependent ribonuclease (RNase L). Transgenic tobacco plants expressing these two enzyme activities were produced by crossing the transgenic plants expressing RNase L with those expressing 2-5Aase. The double transgenic plants showed complete resistance against cucumber mosaic virus (CMV), infection with necrotic spots only forming on the virus-inoculated leaf. On the other hand, although plants inoculated with potato virus Y (PVY) formed necrotic spots on the inoculated leaf and virus amplification could not be detected, all plants died within 20 days of inoculation. The transgenic tobacco plants expressing either 2-5Aase or RNase L activity showed typical disease symptoms with CMV- or PVY-inoculation. These results suggest that the introduced 2-5A system is activated in tobacco cells by dsRNA, the replicating intermediates of RNA viruses, leading to death of the host cells, which has not been observed in mammalian cells.

Transgenic potato expressing a double-stranded RNA-specific ribonuclease is resistant to potato spindle tuber viroid.

We have produced transgenic potato lines expressing the yeast-derived double-stranded RNA-specific ribonuclease pac1. Five lines of pac1 potato (Solanum tuberosum L., cultivar Russet Burbank) challenged with potato spindle tuber viroid (PSTVd) suppressed PSTVd infection and accumulation. All of the progeny potato tubers produced by resistant plants were also free of PSTVd. Because the pac1 gene product digested PSTVd in vitro, double-stranded regions in PSTVd molecule and/or replicative intermediates may be targeted by pac1 gene product in the transgenic potato plant.

Manipulation of human minichromosomes to carry greater than megabase-sized chromosome inserts.

For introducing regions of human chromosomes greater than a megabase into cells or animals, we have developed a chromosome-cloning system in which defined regions of human chromosomes can be cloned into a stable human minichromosome vector in homologous recombination-proficient chicken DT40 cells. The stable minichromosome vector allowed a 10 Mb-sized region of the mitotically unstable human chromosome 22 to be stably maintained in mouse embryonic stem (ES) cells, and in mice. Furthermore, we demonstrated functional expression of human genes from the HAC in mice. This study describes a stable cloning and expression system for greater than megabase-sized regions of human chromosomes.

8-1A, a human monoclonal antibody that reacts with intact human chorionic gonadotropin.

The incidence of choriocarcinoma has decreased over time and therapeutic results have improved about 90% complete remission in patients without extensive metastasis. However, some choriocarcinomas metastasize to other organs and show resistance to chemotherapy, having a poor prognosis despite multidisciplinary treatment. Better methods of early diagnosis for recurrence or micrometastasis, and treatment against cases with intractable gestational trophoblastic neoplasia (GTN) are needed to improve the prognosis. Human chorionic gonadotropin (hCG) is a glycoprotein hormone composed of two dissimilar subunits and a tumor marker to make a diagnosis and monitor therapeutic effect in GTN. Even when hCG levels in the serum become too low to measure with the hCG beta-CTP system which is the most sensitive assay, there are estimated to be approximately 10,000 trophoblastic cells in the body. Residual trophoblast cells may cause symptoms such as bleeding or undergo malignant transformation to choriocarcinoma. Since most monoclonal antibodies developed so far are murine, administration creates human anti-mouse antibodies, resulting in clinical failure. More recent mouse/human chimeric antibodies or humanized antibodies still possess substantial immunogenicity that makes repeated administration difficult. In the present study, KM mice that can produce completely human monoclonal antibodies were used to prepare hCG-specific human monoclonal antibody. This yielded 8-1A, a human monoclonal antibody capable of reacting with intact hCG. In the future, new diagnostic techniques and treatments for chorionic diseases may be developed using this kind of human monoclonal antibody.

[Bronchoplasty without lung resection for the complete transection of the left main bronchus due to blunt trauma].

Blunt bronchial injury is rare but crucial injury. A 17-year-old female was admitted due to traumatic injury. She was diagnosed with bilateral lung contusion, multiple rib fractures, spleen damage and the suspicion about the complete transection of the left main bronchus on X-ray and computed tomography (CT). She was brought to our hospital at 30 hours later from injury. Bronchoscopy revealed the complete transection and the edema of the left main bronchus. She underwent a resection of the disrupted portion and end-to-end anastomosis of left main bronchus without lung resection. We should be an immediate and accurate diagnosis of tracheobronchial disruption by X-ray, CT and bronchoscopy whenever we evaluate patients with blunt chest trauma.

Human normal hepatocytes are susceptible to apoptosis signal mediated by both TRAIL-R1 and TRAIL-R2.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) triggers apoptosis in tumor cells without toxicity to normal cells, but some recombinant versions of TRAIL caused hepatocyte death. We generated fully human monoclonal antibodies (mAbs) that bind specifically to TRAIL receptor 1 (TRAIL-R1) and TRAIL receptor 2 (TRAIL-R2), which mediate apoptosis signal when they ligate with TRAIL, to investigate the contribution of each receptor to induce tumor cell apoptosis and hepatocyte toxicity. All of mAbs to TRAIL-R1 and TRAIL-R2 induced cell death in several cancer cell lines susceptible to TRAIL but not in human umbilical vein endothelial cells in vitro. Both anti-TRAIL-R1 mAbs and anti-TRAIL-R2mAbs also caused cell death in hepatocytes. However, a subset of mAbs to TRAIL-R2, which was characterized by the TRAIL blocking activity, did not show strong hepatocyte toxicity. These results indicate that human normal hepatocytes are susceptible to both TRAIL-R1- and TRAIL-R2-mediated apoptosis signal. Cell Death and Differentiation (2004) 11, 203-207. doi:10.1038/sj.cdd.4401331 Published online 24 October 2003

[Primary lung cancer incidentally diagnosed in lung biopsy for diffuse pulmonary disease].

UNLABELLED: We here presented 2 cases of interstitial pneumonia with lung adenocarcinoma incidentally diagnosed by partially resected lung for diffuse pulmonary disease. CASE 1: A 78-year-old female was admitted to the hospital complaining of productive cough and general fatigue. The chest computed tomography (CT) revealed diffuse honey comb pattern in bilateral lung field especially in the right lower lung. Video-assisted thoracoscopic lung biopsy was performed and was diagnosed as diffuse spreading well differentiated adenocarcinoma. CASE 2: A 59-year-old male was admitted to the hospital complaining of dyspnea and general fatigue. The chest X-ray revealed right pneumothorax and chest CT revealed diffuse honey comb pattern and bullae in bilateral lung field and fibrous tumor-like lesion in the right middle lung. Video-assisted thoracoscopic lung biopsy was performed and was diagnosed as pulmonary fibrosis with papillary adenocarcinoma. CONCLUSION: It is important to examine carefully the specimen obtained from thoracoscopic lung biopsy even if interstitial pneumonia is strongly suspected.

Monoclonal antibodies to choline acetyltransferase of rat brain.

Monoclonal antibodies to rat brain choline acetyltransferase were produced by the hybridoma technique. Two stable cell lines, Ab-57 and Ab-60, secreted immunoglobulin of subclass IgG1. The monoclonal antibodies bound to choline acetyltransferase without blocking catalytic activity. Affinity of Ab-57 was 100-times higher than that of Ab-60. Both antibodies bound to the rat enzyme in a mutually exclusive fashion. The antibodies showed cross-species reactivity with choline acetyltransferase from several mammalian brains.

Resistance against multiple plant viruses in plants mediated by a double stranded-RNA specific ribonuclease.

Many plant viruses have single-stranded RNAs as their genomes. During the course of replication, genomic RNAs and their complementary template RNAs are likely to form double-stranded (ds) RNA at least transiently. To attack replication intermediates of many plant RNA viruses specifically, we introduced a yeast-derived ds-RNA specific RNase gene called pac 1 into plants. The transformed plants showed a decrease in lesion numbers when they were challenged with tomato mosaic virus, and a delay in the appearance of symptoms when inoculated with cucumber mosaic virus or potato virus Y.