Carl G. Hellerqvist

Quantified Color Doppler Sonography of Tumor Vascularity in an Animal Model

This study was designed to evaluate the accuracy of a system to quantitate tumor vascularity with amplitude (power) color Doppler sonography two‐ and three‐dimensionally. The vascularity of 20 transplanted murine tumors was determined with quantitated amplitude color Doppler sonography both two‐ and three‐dimensionally and compared to tumor vascularity estimated by histologic examination. Serial examinations were performed 15, 30, 45, and 60 min after the injection of the exotoxin CM‐101 and saline solution to assess changes in tumor vascularity. Three‐dimensional amplitude color Doppler sonography best depicted the overall vascularity of tumor when compared to histologic estimation of vessel density. However, neither two‐ nor three‐dimensional amplitude color power angiography correlated well to the microvessel count, probably a reflection of the difference in the method for vessel quantification using sonographic versus histologic techniques. Three‐dimensional amplitude Doppler sonography correlated better with counts of large vessels (> 100 microm) as opposed to small vessels (> 15 microm). Time‐activity curves showed no difference in tumor flow at the times measured in the experimental group injected with CM‐101 or when compared to saline solutions in either the peripheral or central portions of the tumor. This three‐dimensional amplitude color Doppler sonographic system affords global quantification of tumor vascularity and flow that may, in turn, be useful in determining the probability of malignancy (by determination of branching patterns and vessel regularity) or tumor response or both to treatment.

Quantification of tumor vascularity and flow with amplitude color Doppler sonography in an experimental model: preliminary results.

This study was designed to evaluate a system to quantitate vascularity and tumor blood flow with amplitude (power) color Doppler sonography. The vascularity of nine transplanted murine tumors was determined with quantitated amplitude color Doppler sonography and compared to tumor vascularity estimated by histologic examination. The system used seemed to provide an accurate depiction of the vascularity of tumor vis‐à‐vis histologic estimation of vessel density (r = 0.80). Time‐activity curves showed greater flow in the experimental group injected with an exotoxin than in the group injected with saline solution. Vascular density quantification with amplitude color Doppler sonography also was more accurate when an intravascular agent (such as an exotoxin) was used than when saline infusions were given. This quantification scheme may allow the development of a system to assess the probability of malignancy and to monitor tumor response to treatment on the basis of the vascularity of the mass.

Structural studies of the common-core polysaccharide of the cell-wall lipopolysaccharide from Salmonella typhimurium

Abstract The structure of the common-core polysaccharide connecting the O-antigenic side-chains to the lipid part in the Salmonella typhimurium lipopolysacc

Studies on Group B β-Hemolytic Streptococcus. II. Effects on Pulmonary Hemodynamics and Vascular Permeability in Unanesthetized Sheep

Summary: To study the effects of Group B β-hemolytic Streptococcus on the pulmonary circulation and lung fluid balance, live and heat-killed bacteria and their toxin were infused into an awake sheep lung-lymph preparation. In every case, the response was biphasic; there was an initial period of marked pulmonary hypertension and high flow of protein-poor lymph associated with tachypnea, chills, and fever. A second phase followed during which pulmonary vascular pressures returned to near baseline and remained stable, but lymph flow remained high. The changes seen in the initial phase resemble the previously reported response to mechanically increased pulmonary vascular pressure and suggest that the increase in fluid filtration is secondary to increased microvascular pressure. During the second phase after toxin infusion, the increase in lung lymph flow was paralleled by an increase in lymph protein clearance. This cannot be accounted for by the hemodynamic changes alone and suggests that the permeability of lung microvascular walls to protein was increased. It is concluded that group B β-hemolytic streptococcal toxin in the sheep model causes pulmonary hypertension and increased pulmonary vascular permeability.Speculation: The effects caused by group G β-hemolytic Streptococcus toxin on the pulmonary circulation may be relevant to the pathogenesis of the respiratory distress and shock seen in newborns with this infection. Further understanding of the effects of this toxin could provide means for therapeutic intervention.

Antitumor effects of GBS toxin: a polysaccharide exotoxin from group B beta-hemolytic streptococcus.

A group B streptococcus (GBS) isolated from human neonates diagnosed with sepsis and respiratory distress (“early-onset disease”) produces a polysaccharide exotoxin (GBS toxin) that, when infused in sheep, causes lung pathophysiology similar to that seen in humans. Histological studies have demonstrated that GBS toxin induces a strong inflammatory response in the lung, with pulmonary sequestration of granulocytes and extensive capillary endothelial damage. The susceptibility of humans to GBS toxin is age-dependent and limited to about 4 days after birth. It is rarely evident thereafter. This suggests that the binding of GBS toxin to the target endothelium occurs via specific components in the developing lung endothelial cells of the newborn that are later lost. We report here that GBS toxin can also bind to developing endothelium associated with neoplasia and induce an inflammatory response. GBS toxin was shown by immunohistochemistry to bind to capillary endothelium of human large-cell carcinomas. In nude mice bearing human tumor xenografts, intravenously administered GBS toxin caused tumor necrosis and hemorrhagic lesions, and substantially inhibited the rate of growth of the tumors. In BALB/c mice bearing Madison lung tumors, GBS toxin induced an inflammatory response resulting in marked changes in tumor morphology, including vasodilation, endothelial and tumor cell necrosis, invasion of lymphocytes and macrophages, and capillary thrombosis. In these tumor models, no evidence of toxicity to the vasculature of other tissues was observed. The reported pathophysiology of GBS in human neonates, the lack of disease in non-neonates colonized with GBS, and these results suggest that GBS toxin may have potential as a well tolerated agent in cancer therapy of some human tumors.

Pulmonary hemodynamic and ultrastructural changes associated with Group B streptococcal toxemia in adult sheep and newborn lambs.

Summary: A toxin isolated from Group B β-hemolytic streptococci, Type III was infused into adult sheep and newborn lambs. A twophased reaction was observed. There was an initial phase of pulmonary hypertension and high flow of protein-poor lymph. This was followed by a second phase when pressures returned to baseline but lymph flow remained twice the baseline values and protein concentration in lymph increased. During the second phase there was a significant increase in lymph protein clearance, suggestive of increased microvascular permeability to protein. The absolute granulocyte count decreased to 10% of baseline values by 60 min after the infusion, and was followed by a variable return to baseline. The sheep with the largest changes in protein clearance were those who had the slowest return to baseline values. Pathologic examination of lung tissue revealed there was capillary dilation, interstitial edema, and large numbers of granulocytes in the lungs. The basement membranes of both capillaries and arterioles showed disruption and widening, along with fragmentation of the internal elastic membrane. This study provides morphologic and physiologic evidence of increased pulmonary vascular permeability after injection of streptococcal toxin associated with granulocyte trapping in the lung. We postulate that granulocytes may be involved as mediators of the pulmonary vascular injury.

Studies on group B beta-hemolytic Streptococcus. I. Isolation and partial characterization of an extracellular toxin.

Summary: To initiate an investigation into the biochemistry and mechanism of group B β-hemolytic Streptococcus virulence, bacterial cultures grown in suspension were centrifuged, and the bacteria and media were subjected to extensive fractionation. Each fraction was assayed for physiologic activity by repeated intravenous infusion into adult unanesthetized sheep. Pulmonary artery pressure, arterial Po2, and rectal temperature were monitored. The media fraction, but not the bacteria, contained a component (molecular weight, 2 X 106) composed of 84% carbohydrate and 16% protein with physiological activity. Two mg quantities, when infused, caused the pulmonary artery pressure to increase 100%, Po2 to decrease by 20% and chills and fever. The active component could be degraded by hot phenol-water extraction into a pure polysaccharide (molecular weight, 200,000). This lower molecular weight polysaccharide retained the identical physiologic properties when infused in the sheep. The degraded component precipitated with group B-specific antiserum.This study demonstrates that, in the sheep, a pure polysaccharide is the physiologically active part of a high-molecular-weight toxin synthesized by group B β-hemolytic Streptococcus type III and that this component has a different carbohydrate composition from the group B capsular antigen.Speculation: The clinical syndrome associated with group B β-hemolytic Streptococci in early onset disease is caused by the interactions of an extracellular bacterial component and a specific target tissue in the infected infant.

The repeating sequence of the capsular polysaccharide of Staphylococcus aureus M

The anomeric configuration of the sugar residues of the capsular polysaccharide antigen of Staphylococcus aureus M were established by 13C-n.m.r. spectroscopy, and the linkage positions by g.l.c.-m.s. after methylation, indicating a leads to 4)-O-(2-acetamido-2-deoxy-alpha-D-galactopyranosyluronic acid)-(1 leads to 4)-O-(2-acetamido-2-deoxy-alpha-D-galactopyranosyluronic acid)-(1 leads to 3)-O-(2-acetamido-2-deoxy-alpha-D-fucopyranosyl)-(1 leads to repeating unit. A taurine residue is linked by an amide bond, on the average, to every fourth 2-acetamido-2-deoxy-D-galactopyranosyluronic acid residue.

CM101, a polysaccharide antitumor agent, does not inhibit wound healing in murine models

CM101 (previously called GBS toxin), a new anticancer polysaccharide that induces inflammatory reactions in neovasculature of tumors, does not cause similar reactions in neovasculature of healing wounds. It appears that treatment with CM101 will not interfere with normal wound healing in cancer patients.

Acute inflammatory changes in subcutaneous microtumors in the ears of mice induced by intravenous CM101 (GBS toxin)

CM101, a bacterial polysaccharide derived from group B streptococcus, induces pronounced inflammatory changes in and around tumor blood vessels 60 min after i. v. injection. A technique has been developed for implanting small numbers of tumor cells in the ear skin of mice. This allows macroscopic examination of the tumor and its supporting blood vessels as it reaches the 10000 cell size and greater. Treatments can be monitored in this model for effects on small “metastatic-like” tumor nodules by direct observation and by histological examination. Inflammatory changes were indicated by increased numbers of polymorphonuclear leukocytes (PMN) adjacent to and marginating within thin-walled blood vessels and within the tumor tissue. PMN were seen in the process of migrating through venules and enlarged capillaries, each with prominent endothelial cells. Tumor morphology was variable with evidence of occasional single necrotic cells. This contrasted with tumors in ears of dextran-treated or untreated mice, which had uniform tumor morphology, and acute inflammatory cells were rarely present.