Cytoadherence characteristics of Plasmodium falciparum isolates in Thailand using an in vitro human lung endothelial cells model

Using an in vitro model of human lung endothelial cells, we studied different characteristics of Plasmodium falciparum isolates as potential factors for malaria severity in 2 Thai patient groups: 27 with complicated malaria and 42 with uncomplicated malaria. In regard to binding properties, no association existed between cytoadherence and rosette phenotypes (P = 0.1) and hypothrombocytemia increased the cytoadherence level (P = 0.007). Cytoadherence was significantly associated with malaria severity (P = 0.05) in contrast to rosette formation (P = 0.9). Intercellular adhesion molecule-1 and chondroitin-4-sulfate were major receptors of cytoadherence in those with complicated malaria compared with those with uncomplicated malaria (P < 10(-4)). Chondroitin-4-sulfate could act as a putative receptor for malaria complications in non-pregnant women. CD36 was the main receptor in patients with uncomplicated malaria (P < 10(-3)). Vascular cell adhesion molecule-1 and E-selectin played a minor role in 2 groups (P = 0.6). Qinghaosu derivatives were more efficient than other antimalarial drugs, but a positive correlation was observed between the 50% inhibitory concentrations of halofantrine and quinine and the number of adhesive parasitized red blood cells, suggesting their influence on cytoadherence.     

Cytomegalovirus and Cryptosporidium infections in AIDS: a necropsy study

A case of coinfection of cytomegalovirus (CMV) and Cryptosporidium in an AIDS patient is reported. Chronic diarrhea was the presenting symptom. Etiologic agents were diagnosed only at postmortem evaluation. CMV intranuclear inclusions were seen in the terminal ileum, colon and vermiform appendix. Cryptosporidium oocysts were also present in the intestinal brush border of the colon. Improvement of diagnostic procedures such as colonic biopsy and the use of appropriate staining procedure for AIDS patients with diarrhea can help identify the cause of illness.     

Cytokines associated with pathology in the brain tissue of fatal malaria

Cytoadherence of Plasmodium falciparum-infected erythrocytes to the brain microvascular endothelial cells is believed to be an important cause of circulatory blockage in cerebral malaria. Cytokines released during acute infection may activate brain endothelial cells leading to increased binding of infected erythrocytes in the brain and reduced cerebral blood flow. This effect may be direct and more potent with the tissue-localized cytokines in the brain. In order to establish this relationship, brain tissues of cerebral and noncerebral malaria were compared. The most prominent histopathologic changes in the brain included edema, neuronal degeneration, ring hemorrhage, and percentage of parasitized erythrocytes sequestration were observed in cerebral malaria. Immunohistochemical staining of the brain sections demonstrated that tissue-localized TNF-alpha, IFN-gamma, IL-I1B, and IL-10 were associated with the histopathology. However, IL-4 was the only cytokine presented at moderate level in the brain tissue of noncerebral malaria which histopathology was the least. No tissue-localized cytokine was observed in the brain of P. vivax infection or of the car accident control cases.     

Functional characterization and target validation of alternative complex I of Plasmodium falciparum mitochondria

This study reports on the first characterization of the alternative NADH:dehydrogenase (also known as alternative complex I or type II NADH:dehydrogenase) of the human malaria parasite Plasmodium falciparum, known as PfNDH2. PfNDH2 was shown to actively oxidize NADH in the presence of quinone electron acceptors CoQ(1) and decylubiquinone with an apparent K(m) for NADH of approximately 17 and 5 muM, respectively. The inhibitory profile of PfNDH2 revealed that the enzyme activity was insensitive to rotenone, consistent with recent genomic data indicating the absence of the canonical NADH:dehydrogenase enzyme. PfNDH2 activity was sensitive to diphenylene iodonium chloride and diphenyl iodonium chloride, known inhibitors of alternative NADH:dehydrogenases. Spatiotemporal confocal imaging of parasite mitochondria revealed that loss of PfNDH2 function provoked a collapse of mitochondrial transmembrane potential (Psi(m)), leading to parasite death. As with other alternative NADH:dehydrogenases, PfNDH2 lacks transmembrane domains in its protein structure, and therefore, it is proposed that this enzyme is not directly involved in mitochondrial transmembrane proton pumping. Rather, the enzyme provides reducing equivalents for downstream proton-pumping enzyme complexes. As inhibition of PfNDH2 leads to a depolarization of mitochondrial Psi(m), this enzyme is likely to be a critical component of the electron transport chain (ETC). This notion is further supported by proof-of-concept experiments revealing that targeting the ETC's Q-cycle by inhibition of both PfNDH2 and the bc(1) complex is highly synergistic. The potential of targeting PfNDH2 as a chemotherapeutic strategy for drug development is discussed.     

Placebo-controlled double-blind cross-over study of botulinum A toxin in hemifacial spasm

The evaluation of the efficacy of botulinum A toxin injection for hemifacial spasm has never previously been done in a double-blind study in spite of its use as a treatment. We thus conducted a double-blind cross-over study of botulinum A toxin use in hemifacial spasm in 55 patients at Siriraj Hospital, Mahidol University, Bangkok, Thailand. Thirteen patients decided to withdraw from the study due to a lack of efficacy, all of them were subsequently found to be in the saline injection group. The remaining 42 patients, in the botulinum A toxin injection (30 mouse units) group, reported the responses as: excellent (34 patients; 80.95%), moderate patients; 2.38%). In contrast, when given the saline injection they reported no excellent outcome, 1 patient (2.38%) with moderate improvement, 5 patients (11.90%) with mild improvement and, 36 patients (85.71%) with no response. Side effects of botulinum toxin injections were found in 14.29% of patients compared with 9.5% of the saline injection group. The side effects of botulinum toxin injection were mild transient facial weakness (7.14%), local pain (4.76%) and excessive lacrimation (2.38%).

We concluded that botulinum A toxin injection was a simple and effective out-patient treatment for the management of hemifacial spasm.     

A clinical trial of combination of artesunate and mefloquine in the treatment of acute uncomplicated falciparum malaria: a short and practical regimen

The difficulties in treating drug-resistant falciparum malaria in Thailand are compounded by the necessity of giving antimalarials over long periods of time. The resultant fall in patient compliance not only lowers cure rates but also predisposes to the further spread of drug-resistance. Sequential treatment with artesunate given over 5 days followed by mefloquine produced 100% cure rates in previous study, but might not be a suitable regimen for field treatment. We conducted a clinical trial of a combination of artesunate and mefloquine given twice daily for 2 days in 150 patients with acute uncomplicated falciparum malaria. The dose of artesunate (200 mg) and mefloquine (312.5 mg) were given simultaneously in a separate package. All patients were admitted to a hospital in Bangkok for 28 days to exclude re-infection and monitor the possible adverse effects. One hundred and thirty patients completed the study with 28 days follow up. Twenty patients (13%) left the hospital prior to completion of follow-up for reasons unrelated to their treatment. Cure rate was 97% (126/130). There were no RII or RIII failures and all four patients with treatment failures were of the RI type. The mean parasite clearance time and fever clearance time were 46.4 and 42.5 hours, respectively. All patients were tolerated the combination drugs well and there were no serious toxic adverse reactions. The results indicate that combination of artesunate and mefloquine given twice daily for 2 days is effective and well tolerated in patients with acute, uncomplicated falciparum malaria and suitable as an alternative treatment for multidrug resistant falciparum malaria.     

Enhanced expression of Fas and FasL modulates apoptosis in the lungs of severe P. falciparum malaria patients with pulmonary edema

Apoptosis mediated by Fas/FasL has been implicated in pulmonary disorders. However, little is known about the relationship between Fas and FasL in the process of lung injury during malaria infection. Paraffin-embedded lung tissues from malaria patients were divided into two groups: those with pulmonary edema (PE) and those without pulmonary edema (non-PE). Normal lung tissues were used as the control group. Cellular expression of Fas, FasL, and the markers of apoptotic caspases, including cleaved caspase-3 and cleaved caspase-8 in the lung tissues were investigated by the immunohistochemistry (IHC) method. Semi-quantitative analysis of IHC staining revealed that cellular expression of Fas, FasL, cleaved caspase-8, and cleaved caspase-3 were significantly increased in the lungs of patients with PE compared with the lungs of patients with non-PE and control groups (all P < 0.05). In addition, significant positive correlations were obtained between Fas and apoptosis (r_s = 0.937, P < 0.001) and FasL and apoptosis (r_s = 0.808, P < 0.001). Significant positive correlations were found between Fas and FasL expression (r_s = 0.827, P < 0.001) and between cleaved caspase-8 and cleaved caspase-3 expression (r_s = 0.823, P < 0.001), which suggests that Fas-dependent initiator and effector caspases, including cleaved caspase-8 and caspase-3, are necessary for inducing apoptosis in the lungs of patients with severe P. falciparum malaria. The Fas/FasL system and downstream activation of caspases are important mediators of apoptosis and may be involved in the pathogenesis of pulmonary edema in severe P. falciparum malaria patients. The proper regulation of the Fas/FasL pathway can be a potential treatment for pulmonary complications in falciparum malaria patients.     

Randomized dose-ranging study of the safety and efficacy of WR 238605 (Tafenoquine) in the prevention of relapse of Plasmodium vivax malaria in Thailand.

WR 238605 is an 8-aminoquinoline developed for the radical cure of Plasmodium vivax. Forty-four P. vivax-infected patients were randomly assigned to 1 of 4 treatment regimens: 3 groups received a blood schizonticidal dose of chloroquine followed by WR 238605: group A (n=15) received 300 mg daily for 7 days; group B (n=11), 500 mg daily for 3 days, repeated 1 week after the initial dose; group C (n=9), 1 dose of 500 mg. A fourth group (D; n=9) received chloroquine only. Among patients who completed 2-6 months of follow-up (n=23), there was 1 relapse in group B (day 120) and 1 in group C (day 112). Among patients treated with chloroquine only, there were 4 relapses (days 40, 43, 49, and 84). WR 238605 was safe, well tolerated, and effective in preventing P. vivax relapse.     

A quantitative ultrastructural study of the liver and the spleen in fatal falciparum malaria

We performed a retrospective study of 25 patients who died of severe falciparum malaria in Thailand and Vietnam using electron microscopy. The aims of the study were: to determine if there was any significant association between parasitized red blood cells (PRBC) sequestered in liver and spleen and particular pre-mortem clinical complications, and to compare the degree of parasite load between the liver and spleen within the same patients. PRBC sequestrations in each organ were compared with the pre-mortem parasitemia, to calculate the sequestration index (S.I.). The S.I. showed that the degree of PRBC sequestration in the spleen was higher than the liver (S.I. median = 3.13, 0.87, respectively) (p < 0.05). The results of quantitative ultrastructural study showed a significantly high parasite load in the liver of patients with jaundice, hepatomegaly and liver enzyme elevation (p < 0.05). We found a significant correlation between PRBC sequestration in the liver and a high serum bilirubin level, a high aspartate aminotransferase (AST) level and an increase in the size of the liver (Spearman's correlation coefficient = 0.688, 0.572, 0.736, respectively). Furthermore, a higher parasite load was found in the liver of patients with acute renal failure (ARF) compared to patients without ARF (p < 0.05). These findings suggest that PRBC sequestration in the liver is quantitatively associated with pre-mortem hepatic dysfunction and renal impairment. There was no significant difference between splenomegaly and PRBC sequestration. The size of a palpable spleen was not correlated with parasite load in the spleen. When ultrastructural features were compared between the two reticuloendothelial organs, we found that the spleen had more PRBC and phagocytes than the liver. The spleen of non-cerebral malaria (NCM) patients had more phagocytes than cerebral malaria (CM) patients. This observation reveals that the spleen plays a major role in malaria parasite clearance, and is associated with host defence mechanisms against malaria.