Antitumor effect of 5-aminolevulinic acid-mediated photodynamic therapy can be enhanced by the use of a low dose of photofrin in human tumor xenografts.

Practically all of the exogenous photosensitizers used for clinical photodynamic therapy (PDT) target mainly vasculature. Although effective in tumor destruction, they also, unavoidably, induce phototoxicity of normal tissues. Porphyrins synthesized endogenously from 5-aminolevulinic acid (ALA) accumulate within cells. Tumor eradication would be more efficient if both cellular components and vascular stroma of a tumor could be targeted. Thus, PDT with a mixture of ALA and Photofrin (Pf, a vessel-targeted sensitizer) may simultaneously destroy the two elements. Using chemical extraction assays, pharmacokinetics of ALA and ALA-induced porphyrins were studied in the plasma and tumors of nude mice bearing human WiDr and KM20L2 colonic carcinomas after an i.p. injection of 250 mg/kg body weight of ALA. Subsequently, PDT efficacy of the two tumor models with ALA, Pf, or with the two drugs in combination was evaluated. The phototoxic effects on tumor cells in vitro with the combined drugs was also determined. Moreover, histological and ultrastructural alterations of the treated tumors were investigated, and tumor cell clonogenicity was assessed as a function of time after in vivo PDT using an in vitro colony formation assay. Finally, the photosensitivity of normal skin tissue treated according to various protocols was compared. The amounts of ALA peaked at 0.5 h after administration in both plasma and WiDr tumor. The rates of ALA clearance seemed to follow a one-compartment model with half-lives of approximately 18 and 58 min in the plasma and tumor, respectively. About 100 and 60 times higher concentrations of ALA were needed to induce a given concentration of porphyrins in the plasma and tumor, respectively, although the plasma porphyrins may not only be released from blood cells but also from other organs. Similar kinetics of distribution patterns of ALA- and ALA methylester-induced porphyrins were found in the plasma and tumors, and the elimination rates were consistent with a two-compartment model. ALA induced much more porphyrins than ALA methylester in both plasma and tumors. Tumors PDT-treated with ALA plus Pf at a low dose (1 mg/kg) grew significantly more slowly than those treated with either of the drugs in both WiDr and KM20L2 models. However, the enhanced antitumor effect was not found in the tumor cells under in vitro conditions. Morphological studies demonstrated that PDT with the combined regimen resulted in necrosis of neoplastic cells and severe disruption of tumor microvasculature. This was supported by the findings obtained from the studies of in vivo PDT and in vitro clonogenic assay that showed a progressive reduction in tumor cell viability with times following PDT. Such a combined PDT protocol did not induce any phototoxicity in normal skin tissue. These data indicate that targeting both neoplastic cells and stroma with ALA and Pf (a low dose) can potentiate antitumor PDT effect with no risk of prolonged skin photosensitivity.

A piglet survival model of posthypoxic encephalopathy.

The aim of this study was to produce a neonatal piglet model which, avoiding vessel ligation, exposed the whole animal to hypoxia and produced dose-dependent clinical encephalopathy and neuropathologic damage similar to that seen after birth asphyxia. Twenty-three piglets were halothane-anesthetized. Hypoxia was induced in 19 piglets by reducing the fractional concentration of inspired oxygen (FiO2) to the maximum concentration at which the EEG amplitude was below 7 microV (low amplitude) for 17-55 min. There were transient increases in Fio2 to correct bradycardia and hypotension. Posthypoxia, the piglets were extubated when breathing was stable. Four were sham-treated controls. We aimed at 72-h survival; seven died prematurely due to posthypoxic complications. EEG and a videotaped itemized neurologic assessment were recorded regularly. We found that 95% of the animals showed neuropathologic damage. The duration of low amplitude EEG during the insult and the arterial pH at the end of the insult correlated with cortical/white matter damage; r = 0.75 and 0.81, respectively. Early postinsult EEG background amplitude (r = 0.86 at 3 h) and neurologic score (r = 0.79 at 8 h) correlated with neuropathology. Epileptic seizures in seven animals were always associated with severe neuropathologic damage. We conclude that EEG-controlled hypoxia and subsequent intensive care enabled the animals to survive with an encephalopathy which correlated with the cerebral hypoxic insult. The encephalopathy was clinically, electrophysiologically, and neuropathologically similar to that in the asphyxiated term infant. This model is suitable for examining mechanisms of damage and evaluation of potential protective therapies after birth asphyxia.

Haemoglobin is inversely related to plasma lactate and heart rate in the newborn piglet.

It has been hypothesized that an increased level of plasma lactate (La), the product of anaerobic glycolysis, may identify infants with anaemia and early tissue hypoxia. In the newborn piglet, a frequently used model in experimental neonatology, there is a substantial decrease in haemoglobin (Hb) during the first days of life. We have investigated whether a low level of Hb is associated with an increased level of La in the newborn piglet. Arterial blood gas, Hb, glucose, La, and continuous blood pressure and heart rate (HR) recordings were obtained from 66 newborn piglets. In 4 additional piglets recordings were obtained before isovolemic anaemia, during isovolemic anaemia, and after retransfusion of packed red cells. A low concentration of Hb in the newborn piglet correlates with an increased level of La as well as with an increased HR. A reduction of Hb causes an increase in La and in HR. We hypothesize that increased La and increased HR are caused by mild hypoxia due to normal, low Hb levels in the apparently healthy, resting newborn piglet.

Posthypoxic cooling of neonatal rats provides protection against brain injury.

AIM: To determine whether moderate hypothermia, applied after a hypoxic-ischaemic insult in neonatal rats, reduces cerebral damage. METHOD: Unilateral hypoxic-ischaemic brain damage was induced in 7 day old rats by left carotid ligation, followed by 120 minutes of normothermic exposure to 8% O2, followed by random selection to three hours of hypothermia (rectal temperature, mean (SD), 32.5 (0.4) degrees C) or normothermia (38.3 (0.4) degrees C). One hundred and one animals were used for brain temperature or blood chemistry studies and 24 for survival studies (7 days) with neuropathology, including cell counting as outcome measures. RESULTS: Thirty sections from each brain were histologically examined with respect to distribution and pattern of damage and given a score from 0 to 4. Animals treated with hypothermia had significantly less damage than normothermic animals (score 0.5 (0.3) vs 1.8 (0.5)). CONCLUSIONS: Posthypoxic hypothermia reduces brain damage in awake, unrestrained 7 day old rats.