Diagnostic ultrasound activation of contrast agent gas bodies induces capillary rupture in mice

Interaction of diagnostic ultrasound with gas bodies produces a useful contrast effect in medical images, but the same interaction also represents a mechanism for bioeffects. Anesthetized hairless mice were scanned by using a 2.5-MHz transducer (610-ns pulses with 3.6-kHz repetition frequency and 61-Hz frame rate) after injection of Optison and Evans blue dye. Petechial hemorrhages (PHs) in intestine and abdominal muscle were counted 15 min after exposure to characterize capillary rupture, and Evans blue extravasation was evaluated in samples of muscle tissue. For 5 ml⋅kg-1 contrast agent and exposure to 10 alternating 10-s on and off periods, PH counts in muscle were approximately proportional to the square of peak negative pressure amplitude and were statistically significant above 0.64 MPa. PH counts in intestine and Evans blue extravasation into muscle tissue were significant above 1.0 MPa. The PH effect in muscle was proportional to contrast dose and was statistically significant for the lowest dose of 0.05 ml⋅kg-1. The effects decreased nearly to sham levels if the exposure was delayed 5 min. The PH effect in abdominal muscle was significant and statistically indistinguishable for uninterrupted 100-s exposure, 10-s exposure, 100 scans repeated at 1 Hz, and even for a single scan. The results confirms a previous report of PH induction by diagnostic ultrasound with contrast agent in mammalian skeletal muscle [Skyba, D. M., Price, R. J., Linka, A. Z., Skalak, T. C. & Kaul, S. (1998) Circulation 98, 290–293].

Tolerance of human MSH2+/− lymphoblastoid cells to the methylating agent temozolomide

Members of hereditary nonpolyposis colon cancer (HNPCC) families harboring heterozygous germline mutations in the DNA mismatch repair genes hMSH2 or hMLH1 present with tumors generally two to three decades earlier than individuals with nonfamilial sporadic colon cancer. We searched for phenotypic features that might predispose heterozygous cells from HNPCC kindreds to malignant transformation. hMSH2+/− lymphoblastoid cell lines were found to be on average about 4-fold more tolerant than wild-type cells to killing by the methylating agent temozolomide, a phenotype that is invariably linked with impairment of the mismatch repair system. This finding was associated with an average 2-fold decrease of the steady-state level of hMSH2 protein in hMSH2+/− cell lines. In contrast, hMLH1+/− heterozygous cells were indistinguishable from normal controls in these assays. Thus, despite the fact that HNPCC families harboring mutations in hMSH2 or hMLH1 cannot be distinguished clinically, the early stages of the carcinogenic process in hMSH2 and hMLH1 mutation carriers may be different. Should hMSH2+/− colonocytes and lymphoblasts harbor a similar phenotype, the increased tolerance of the former to DNA-damaging agents present in the human colon may play a key role in the initiation of the carcinogenic process.

A general genetic approach in Escherichia coli for determining the mechanism(s) of action of tumoricidal agents: application to DMP 840, a tumoricidal agent.

We describe here a simple and easily manipulatable Escherichia coli-based genetic system that permits us to identify bacterial gene products that modulate the sensitivity of bacteria to tumoricidal agents, such as DMP 840, a bisnaphthalimide drug. To the extent that the action of these agents is conserved, these studies may expand our understanding agents is conserved, these studies may expand our understanding of how the agents work in mammalian cells. The approach briefly is to use a library of E. coli genes that are overexpressed in a high copy number vector to select bacterial clones that are resistant to the cytotoxic effects of drugs. AtolC bacterial mutant is used to maximize permeability of cells to hydrophobic organic molecules. By using DMP 840 to model the system, we have identified two genes, designated mdaA and mdaB, that impart resistance to DMP 840 when they are expressed at elevated levels. mdaB maps to E. coli map coordinate 66, is located between the parE and parC genes, and encodes a protein of 22 kDa. mdaA maps to E. coli map coordinate 18, is located adjacent to the glutaredoxin (grx) gene, and encodes a protein of 24 kDa. Specific and regulatable overproduction of both of these proteins correlates with DMP 840 resistance. Overproduction of the MdaB protein also imparts resistance to two mammalian topoisomerase inhibitors, Adriamycin and etoposide. In contrast, overproduction of the MdaA protein produces resistance only to Adriamycin. Based on its drug-resistance properties and its location between genes that encode the two subunits of the bacterial topoisomerase IV, we suggest that mdaB acts by modulating topoisomerase IV activity. The location of the mdaA gene adjacent to grx suggests it acts by a drug detoxification mechanism.