The mechanism of inhibition of the sarco/endoplasmic reticulum Ca2+ ATPase by paxilline

Although cis-diamminedichloroplatinum (II) (cisplatin) is a potent anticancer drug, clinical use of this agent is highly limited predominantly because of its strong side effects on the kidney and gastrointestinal tracts. We found that cisplatin impaired respiratory function and DNA of mitochondria in renal proximal tubules and small intestinal mucosal cells, thereby inducing apoptosis of epithelial cells. Cisplatin-induced mitochondrial dysfunction and DNA (mtDNA) injury, lipid peroxidation, and apoptosis of epithelial cells in the kidney and small intestine were strongly inhibited by L-carnitine. However, carnitine had no appreciable effect on the tumoricidal action of cisplatin against cancer cells inoculated in the peritoneal cavity. These results indicate that L-carnitine may have therapeutic potential for inhibiting the side effects of cisplatin and other anticancer agents in the kidney and small intestine.     

6-Gingerol prevents cisplatin-induced acute renal failure in rats

BACKGROUND: Nephrotoxicity is a major complication and a dose limiting factor for cisplatin therapy. Recent evidence suggests that enhanced oxidative stress caused by oxygen-centered free radicals may contribute to the pathogenesis of cisplatin-induced acute renal failure. 6-Gingerol is claimed to be a potent antioxidant. The present study was performed to explore the renoprotective potential of 6-gingerol on cisplatin-induced oxidative stress and renal dysfunction. METHODS: 6-Gingerol in dosages of 12.5, 25, 50 mg/kg was administered 2 days before and 3 days after cisplatin administration. Renal injury was assessed by measuring serum creatinine, blood urea nitrogen, creatinine, urea clearance and serum nitrite levels. Renal oxidative stress was assessed by determining renal malondialdehyde levels, reduced glutathione levels and enzymatic activities of superoxide dismutase and catalase. RESULTS: A single dose of cisplatin resulted in marked renal oxidative and nitrosative stress and significantly deranged renal functions. 6-Gingerol treatment significantly and dose-dependently restored renal functions, reduced lipid peroxidation and enhanced the levels of reduced glutathione and activities of superoxide dismutase and catalase. CONCLUSIONS: The present study demonstrates the renoprotective potential of 6-gingerol against cisplatin-induced oxidative stress and renal dysfunction in rats. Hence, 6-gingerol has a potential to be used as therapeutic adjuvant in cisplatin nephrotoxicity.     

Maternal antioxidant concentrations after uncomplicated pregnancies

Background: To analyse the post-partum concentrations of intra- and extra-cellular blood antioxidants in women with uncomplicated pregnancies.

Methods: Whole blood and plasma thiols, plasma vitamin E and C, serum cholesterol and triglyceride, ferric reducing ability of plasma (FRAP) concentrations were compared between women delivered by caesarean section (n=17) or spontaneous delivery (n=10). A repeated mixed model was used for statistical analysis.

Results: The majority of whole blood thiols increased significantly in both groups the first days post-partum. However, within the caesarean group free cysteine, oxidised cysteine, homocysteine and glutathione and plasma cysteine and homocysteine levels dropped significantly after 24 h, while FRAP levels peaked significantly in this group. Plasma vitamin E levels decreased significantly in both groups within 24 to 48 h after delivery. Independent of the way of delivery whole blood and plasma thiols were significantly increased and vitamin E levels were significantly decreased 3 months post-partum while plasma vitamin C levels and FRAP were unchanged compared to ante-partum levels.

Discussion: Decreased plasma vitamin E levels shortly post-partum are associated with decreased lipid peroxidation. The 24 h post-partum drop of some plasma and whole blood thiols in the caesarean group may be due to prolonged fasting.     

Mitochondrial-targeted antioxidants represent a promising approach for prevention of cisplatin-induced nephropathy

Cisplatin is a widely used antineoplastic agent; however, its major limitation is the development of dose-dependent nephrotoxicity whose precise mechanisms are poorly understood. Here we show not only that mitochondrial dysfunction is a feature of cisplatin nephrotoxicity, but also that targeted delivery of superoxide dismutase mimetics to mitochondria largely prevents the renal effects of cisplatin. Cisplatin induced renal oxidative stress, deterioration of mitochondrial structure and function, an intense inflammatory response, histopathological injury, and renal dysfunction. A single systemic dose of mitochondrially targeted antioxidants, MitoQ or Mito-CP, dose-dependently prevented cisplatin-induced renal dysfunction. Mito-CP also prevented mitochondrial injury and dysfunction, renal inflammation, and tubular injury and apoptosis. Despite being broadly renoprotective against cisplatin, Mito-CP did not diminish cisplatin's antineoplastic effect in a human bladder cancer cell line. Our results highlight the central role of mitochondrially generated oxidants in the pathogenesis of cisplatin nephrotoxicity. Because similar compounds seem to be safe in humans, mitochondrially targeted antioxidants may represent a novel therapeutic approach against cisplatin nephrotoxicity.     

Autophagy delays apoptosis in renal tubular epithelial cells in cisplatin cytotoxicity

One of the major side effects of cisplatin chemotherapy is toxic acute kidney injury due to preferential accumulation of cisplatin in renal proximal tubule epithelial cells and the subsequent injury to these cells. Apoptosis is known as a major mechanism of cisplatin-induced cell death in renal tubular cells. We have also recently demonstrated that autophagy induction is an immediate response of renal tubular epithelial cell exposure to cisplatin. Inhibition of cisplatin-induced autophagy blocks the formation of autophagosomes and enhances cisplatin-induced caspase-3, -6, and -7 activation, nuclear fragmentation, and apoptosis. The switch from autophagy to apoptosis by autophagic inhibitors suggests that autophagy induction was responsible for a pre-apoptotic lag phase observed on exposure of renal tubular cells to cisplatin. Our studies provide evidence that autophagy induction in response to cisplatin mounts an adaptive response that suppresses and delays apoptosis. The beneficial effect of autophagy has a potential clinical significance in minimizing or preventing cisplatin nephrotoxicity.

Addedum to: Yang C, Kaushal V, Shah SV, Kaushal GP. Autophagy and apoptosis are associated in cisplatin injury to renal tubular epithelial cell injury. Am J Physiol Renal Physiol 2008; 294:F777-87.     

Involvement of heme oxygenase as antioxidant defense in soybean nodules

Objective: We have previously demonstrated that the inducible form of heme oxygenase plays a critical role in protecting against oxidative stress in mammals. To gain further insight into the functions of this enzyme in plants, we have tested its activity and expression in soybean nodules subjected to cadmium (Cd) stress.

Materials and methods: Four-weeks-old soybean nodulated plants were treated with different cadmium chloride concentrations (0, 50 and 200 μM) during 48 h. Oxidative stress parameters such as TBARS content, GSH levels and antioxidant enzyme activities were measured as well as heme oxygenase activity and expression. Besides, the effect of biliverdin and Zn-protophorphyrin IX were analized.

Results: Treatment with 200 μM Cd during 48 h caused a 67% increase in TBARS content, whereas GSH decreased 44%, and total superoxide dismutase, gluthatione reductase and guaiacol peroxidase were also inhibited 54, 20 and 60%, respectively. A total of 200 μM Cd produced the overexpression of heme oxygenase-1, as well as a 10-fold enhancement of its activity. Co-administration of biliverdin (10 μM) completely prevented the effects caused by Cd. Treatment with Zn protoporphyrin IX, a strong inhibitor of heme oxygenase, expectedly decreased heme oxygenase-1 activity to half. When the inhibitor was given together with Cd, completely prevented the enzyme induction and oxidative stress parameters were significantly enhanced.

Conclusion: Taking together, these results are indicating that heme oxygenase plays a protective role against oxidative cell damage in soybean nodules.     

Tim-1 promotes cisplatin nephrotoxicity

Nephrotoxicity is a frequent complication of cisplatin-based chemotherapy, in which T cells are known to promote acute kidney injury. In this study, we examined the role of T cell immunoglobulin mucin 1 (Tim-1) in cisplatin-induced acute kidney injury using an inhibitory anti-Tim-1 antibody. Tim-1 acts to modulate T cell responses, but it is also expressed by damaged proximal tubules in the kidney, where it is known as kidney injury molecule-1 (Kim-1). Anti-Tim-1 antibodies attenuated cisplatin nephrotocity, with less histologic damage, improved renal function, and fewer leukocytes infiltrating the kidney compared with control antibody-treated mice. Renal NF-κB activation and apoptosis were reduced, and proinflammatory renal cytokine and chemokine mRNA expression was decreased. Renal Kim-1 expression was reduced, consistent with the diminished kidney injury after anti-Tim-1 antibody treatment. Furthermore, anti-Tim-1 antibodies reduced early systemic CD4+ and CD8+ T cell activation, apoptosis, and cytokine production. To determine whether the protective actions of anti-Tim-1 antibodies were due to effects on renal tubular cells, cisplatin nephrotoxicity was studied in Rag1(-/-) mice. Anti-Tim-1 antibodies did not affect renal dysfunction or histologic damage in Rag1(-/-) mice, showing that the benefits of inhibiting Tim-1 come from T cell effects. As Tim-1 plays an important role in promoting cisplatin nephrotoxicity, inhibiting Tim-1 may be a therapeutic strategy to prevent cisplatin-induced acute kidney injury.     

PSTPIP2 inhibits cisplatin-induced acute kidney injury by suppressing apoptosis of renal tubular epithelial cells

Cisplatin (CP) is an effective chemotherapeutic agent widely used in the treatment of various solid tumours. However, CP nephrotoxicity is an important limitation for CP use; currently, there is no method to ameliorate cisplatin-induced acute kidney injury (AKI). Recently, we identified a specific role of proline–serine–threonine phosphatase-interacting protein 2 (PSTPIP2) in cisplatin-induced AKI. PSTPIP2 was reported to play an important role in a variety of diseases. However, the functions of PSTPIP2 in experimental models of cisplatin-induced AKI have not been extensively studied. The present study demonstrated that cisplatin downregulated the expression of PSTPIP2 in the kidney tissue. Administration of AAV-PSTPIP2 or epithelial cell-specific overexpression of PSTPIP2 reduced cisplatin-induced kidney dysfunction and inhibited apoptosis of renal tubular epithelial cells. Small interfering RNA-based knockdown of PSTPIP2 expression abolished PSTPIP2 regulation of epithelial cell apoptosis in vitro. Histone acetylation may impact gene expression at the epigenetic level, and histone deacetylase (HDAC) inhibitors were reported to prevent cisplatin-induced nephrotoxicity. The UCSC database was used to predict that acetylation of histone H3 at lysine 27 (H3K27ac) induces binding to the PSTPIP2 promoter, and this prediction was validated by a ChIP assay. Interestingly, an HDAC-specific inhibitor (TSA) was sufficient to potently upregulate PSTPIP2 in epithelial cells. Histone acetylation-mediated silencing of PSTPIP2 may contribute to cisplatin nephrotoxicity. PSTPIP2 may serve as a potential therapeutic target in the prevention of cisplatin nephrotoxicity.Subject terms: Cancer, Cancer prevention     

TLR2 protects cisplatin-induced acute kidney injury associated with autophagy via PI3K/Akt signaling pathway

Toll-like receptors (TLRs), which are essential components of the innate immune response, play an important role in acute kidney injury (AKI). Toll-like receptor 2 (TLR2) is constitutively expressed in tubular epithelial cells of the kidney and participates in cisplatin-induced AKI. The autophagy is a dynamic catabolic process that maintains intracellular homeostasis, which is involved in the pathogenesis of AKI. Recent studies demonstrate that PI3K/Akt signaling pathway regulates autophagy in response to various stimuli. Therefore, we propose that cisplatin might activate TLR2, which subsequently phosphorylates PI3K/Akt, leading to enhanced autophagy of renal tubular epithelial cells and protecting cisplatin-induced AKI. We found that TLR2 expression was significantly increased in the kidney after the cisplatin treatment. TLR2-deficient mice exacerbated renal injury in cisplatin-induced AKI, with higher serum creatinine and blood urea nitrogen, more severe morphological injury compared with that of wild-type mice. In vitro, we found that inhibition of TLR2 reduced tubular epithelial cell autophagy after the cisplatin treatment. Mechanistically, TLR2 inhibited autophagy via activating PI3K/Akt signaling pathway in renal tubular epithelial cells after the cisplatin treatment. Take together, these results suggest that TLR2 may protect cisplatin-induced AKI by activating autophagy via PI3K/Akt signaling pathway.     

Carvedilol protects against cisplatin-induced oxidative stress, redox state unbalance and apoptosis in rat kidney mitochondria

Cisplatin is a highly effective chemotherapeutic agent which causes severe nephrotoxicity. Studies have suggested that reactive oxygen species, mainly generated in mitochondria, play a central role in cisplatin-induced renal damage. A wide range of antioxidants have been evaluated as possible protective agents against cisplatin-induced nephrotoxicity; however a safe and efficacious compound has not yet been found. The present study is the first to evaluate the protective potential of carvedilol, a beta-blocker with strong antioxidant properties, against the mitochondrial oxidative stress and apoptosis in kidney of rats treated with cisplatin. The following cisplatin-induced toxic effects were prevented by carvedilol: increased plasmatic levels of creatinine and blood urea nitrogen (BUN); lipid peroxidation, oxidation of cardiolipin; oxidation of protein sulfhydryls; depletion of the non-enzymatic antioxidant defense and increased activity of caspase-3. Carvedilol per se did not present any effect on renal mitochondria. It was concluded that carvedilol prevents mitochondrial dysfunction and renal cell death through the protection against the oxidative stress and redox state unbalance induced by cisplatin. The association of carvedilol to cisplatin chemotherapy was suggested as a possible strategy to minimize the nephrotoxicity induced by this antitumor agent.     

Autophagy delays apoptosis in renal tubular epithelial cells in cisplatin cytotoxicity

One of the major side effects of cisplatin chemotherapy is toxic acute kidney injury due to preferential accumulation of cisplatin in renal proximal tubule epithelial cells and the subsequent injury to these cells. Apoptosis is known as a major mechanism of cisplatin-induced cell death in renal tubular cells. We have also recently demonstrated that autophagy induction is an immediate response of renal tubular epithelial cell exposure to cisplatin. Inhibition of cisplatin-induced autophagy blocks the formation of autophagosomes and enhances cisplatin-induced caspase-3, -6, and -7 activation, nuclear fragmentation and apoptosis. The switch from autophagy to apoptosis by autophagic inhibitors suggests that autophagy induction was responsible for a pre-apoptotic lag phase observed on exposure of renal tubular cells to cisplatin. Our studies provide evidence that autophagy induction in response to cisplatin mounts an adaptive response that suppresses and delays apoptosis. The beneficial effect of autophagy has a potential clinical significance in minimizing or preventing cisplatin nephrotoxicity.     

Capsaicin Ameliorates Cisplatin-Induced Renal Injury through Induction of Heme Oxygenase-1

Cisplatin is one of the most potent chemotherapy agents. However, its use is limited due to its toxicity in normal tissues, including the kidney and ear. In particular, nephrotoxicity induced by cisplatin is closely associated with oxidative stress and inflammation. Heme oxygenase-1 (HO-1), the rate-limiting enzyme in the heme metabolism, has been implicated in a various cellular processes, such as inflammatory injury and anti-oxidant/oxidant homeostasis. Capsaicin is reported to have therapeutic potential in cisplatin-induced renal failures. However, the mechanisms underlying its protective effects on cisplatin-induced nephrotoxicity remain largely unknown. Herein, we demonstrated that administration of capsaicin ameliorates cisplatin-induced renal dysfunction by assessing the levels of serum creatinine and blood urea nitrogen (BUN) as well as tissue histology. In addition, capsaicin treatment attenuates the expression of inflammatory mediators and oxidative stress markers for renal damage. We also found that capsaicin induces HO-1 expression in kidney tissues and HK-2 cells. Notably, the protective effects of capsaicin were completely abrogated by treatment with either the HO inhibitor ZnPP IX or HO-1 knockdown in HK-2 cells. These results suggest that capsaicin has protective effects against cisplatin-induced renal dysfunction through induction of HO-1 as well as inhibition oxidative stress and inflammation.     

Book Reviews

The Biology of Nitric Oxide, vol. 3. Physiological and Clinical Aspects; vol. 4. Enzymology, Biochemistry and Immunology, Portland Press, 1994

Mitochondrial Diseases Editors: L. Ernster, R. Luft and S. Orrenius Publishers: Elsevier

Biothiols in Health and Disease Editors: Lester Packer & Enrique Cadenas Publishers: Marcel Dekker, Inc.     

Infradian dynamics and variability of salivary testosterone in men and women

Background. The dynamics of testosterone levels exhibits several cyclic patterns with various period lengths. Circadian and circannual rhythms of testosterone are known in both genders. Among infradian rhythms only the circalunar cycle in women is widely accepted. In our previous studies we have found a circatrigintan (30 days) and a circavigintan (20 days) cycle in men. Whether cyclic patterns with higher frequencies are present in the dynamics of testosterone levels in men or in women is unknown.

Aim. To analyze the infradian dynamics of salivary testosterone in both genders for the presence of cyclic patterns.

Subjects and methods. Seventeen young and healthy women and 15 men were asked to collect saliva samples during 30 consecutive days. Testosterone levels were determined using radioimmunoassay, Analysis of Rhythmic Variance II (ANORVA II) was used for statistical analysis. Potential period lengths of 3 - 15 days were evaluated.

Results. The dynamics of salivary testosterone showed high intra-individual variability in both genders (coefficient of variation - 28% in women and 26% in men). ANORVA II analysis showed no significant rhythms, although a weak circaseptan cyclic pattern has been found in women.

Discussion. Our results showed no significant infradian cyclic variation with a period between 3 and 15 days. Further studies should concentrate on potential longer periods. Described intra-individual variability of testosterone levels in both genders should be considered in endocrine research.     

Prevention and repair of cerebral ischemia-reperfusion injury by Chinese herbal medicine, Shengmai San, in rats

The protective activity of Shengmai San, a traditional Chinese herbal medicine, was studied in cerebral ischemia-reperfusion injury in rats. Shengmai San consists of three herbal components, Panax Ginseng, Ophiopogon Japonicus and Schisandra Chinensis and is routinely being used for treating coronary heart disease.

When Shengmai San was injected directly into rat duodenum 2 h before cerebral ischemia by bilateral carotid artery occlusion, thiobarbituric acid reactive substance (TBARS) formation during reperfusion following ischemia was almost completely suppressed in the brain. The loss of glutathione perioxidase activity after the ischemia-reperfusion was also effectively prevented by the Shengmai San pre-administration whereas the activity was considerably decreased in the damaged brain.

It was found that Shengmai San also effectively suppressed the TBARS formation even when it was administered after 45 min reperfusion following ischemia, indicating that Shengmai San improves the oxidative damage already established in the brain. Likewise, the decrease of glutathione peroxidase activity was minimized in the damaged brain by the post-administration of Shengmai San.

On the other hand, none of the Shengmai San components were active in protecting the ischemia-reperfusion brain damage when they were independently administered.

These experiments suggest the potential of Shengmai San in both preventive and therapeutic usages for cerebral ischemia-reperfusion injury.     

Book Review

LEGGETT, W. F. Ancient and Medieval Dyes. 5 × 8 in. 96 pp. Cloth. Chemical Publishing Co., Brooklyn, N. Y. 1944. $2.25.

Microtechnic In General. McCARTNEY, J. E. A new immersion oil “polyric”. J. Path. & Bact., 56, 265-6. 1944.

NICKERSON, MARK. A dry ice freezing unit for rotary microtomes. Science, 100, 177-8. 1944.

Dyes And Thedx Biological Uses. BERGEIM, FRANK H., and BRAKER, WILLIAM. Homosulfanilamides. J. Amer. Chem. Soc., 66, 1459. 1944.

CALDWELL, W. T., TYSON, F. T., and LAUER, LOTHAR. Substituted 2-sulfonamido-5-aminopyridines. II. J. Amer. Chem. Soc., 66, 1479. 1944.

JOHNS, C. K. Dye concentration in resazurin tablets. Amer. J. Pub. Health, 34, 955-8. 1944.

SMITH, WINSLOW WHITNEY. Relative sensitivity of different phases of growth curve of Bact. salmonicida to alkaline acriflavine. Proc. Soc. Exp. Biol. & Med., 56, 240-2. 1844.

VAN ARENDONK, A. M., and SHOULE, H. A. Dialkylaminoalkyl derivatives of substituted quinolines and quinaldines. J. Amer. Chem. Soc., 66, 1284. 1944.

WHEELER, KEITH, and DEGERING, E. F. Preparation and properties of certain derivatives of sulfamide. J. Amer. Chem. Soc., 66, 1242. 1944.

Animal Microtechnic. BOARDMAN, EDWARD T. Methods for collecting ticks for study and delineation. J. Parasitology, 30, 57-9. 1944.

DICKIE, MARGARET M. A new differential stain for mouse pituitary. Science, 100, 297-8. 1944.

GOVAN, A. D. TELFORD. Fat staining by Sudan dyes suspended in watery media. J. Path. & Bact., 56, 262-4. 1944.

LILLIE, R. D., and ASHBURN, L. L. Supersaturated solutions of fat stains in dilute isopropanol for demonstration of acute fatty degenerations not shown by Herxheimer technic. Arch. Path., 36, 432. 1943.

MULLEN, J. P. A convenient and rapid method for staining glycogen in paraffin sections with Best's carmine stain. Amer. J. Clin. Path., Tech. Sect., 8, 9-10. 1944.

NYKA, W. A method for staining the rickettsiae of typhns in histological sections. J. Path. & Bact., 56, 264. 1944.

POPPER, HANS, GYORGY, PAUL, and GOLDBLATT, H. Fluorescent material (ceroid) in experimental nutritional cirrhosis. Arch. Path., 37, 161. 1944.

SMALL, C. S., and SCHULTZ, M. A. Sustaining faded tissue sections. Amer. J. Clin. Path., Tech. Sect., 7, 66-7. 1943.

YOFFEY, J. M., and PARNELL, J. The lymphocyte content of rabbit bone marrow. J. Anat., 78, 109-12. 1944.

ZIEGLER, E. E. Hematoxylin-eosin tissue stain. An improved, rapid, and uniform technic. Arch. Path., 37, 68. 1044.

Plant Microtechnic. HAASIS, FERDINAND W. Staining rubber in ground or milled plant tissues. Ind. and Eng. Chem., Anal. Ed., 16, 480. 1944.

PARRIS, G. K. A simple nuclear stain and staining technique for Helminthosporia. Phytopathology, 34, 700. 1944.

Microorganisms. DARZINS, E. Rickettsienstudien. Zentbl. Bakl., Abt. I, Orig., 151, 18-20. 1943.

GOHAR, M. A. A staining method for Corynebacterium diphtheriae. J. Bact., 47, 575. 1944.

GRAY, P. H. H. Two-stain method for direct bacteria count. J. Milk Techn., 6, 76. 1943.     

Protein kinase C-alpha and ERK1/2 mediate mitochondrial dysfunction,decreases in active Na+ transport,and cisplatin-induced apoptosis in renal cells

Initiation of apoptosis by many agents is preceded by mitochondrial dysfunction and depolarization of the mitochondrial inner membrane. Here we demonstrate that, in renal proximal tubular cells (RPTC), cisplatin induces mitochondrial dysfunction associated with hyperpolarization of the mitochondrial membrane and that these events are mediated by protein kinase C (PKC)-alpha and ERK1/2. Cisplatin induced sustained decreases in RPTC respiration, oxidative phosphorylation, and increases in the mitochondrial transmembrane potential (deltaPsi(m)), which were preceded by the inhibition of F(0)F(1)-ATPase and cytochrome c release from the mitochondria, accompanied by caspase-3 activation, and followed by RPTC apoptosis. Cisplatin also decreased active Na+ transport as a result, in part, of the inhibition of Na+/K(+)-ATPase. These changes were preceded by PKC-alpha and ERK1/2 activation. Inhibition of cisplatin-induced PKC-alpha and ERK1/2 activation using Go6976 and PD98059, respectively, abolished increases in deltaPsi(m), diminished decreases in oxidative phosphorylation, active Na+ transport, and decreased caspase-3 activation without blocking cytochrome c release. Caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (zVAD-fmk) did not prevent increases in deltaPsi(m). Furthermore, inhibition of PKC-alpha did not prevent cisplatin-induced ERK1/2 activation. We concluded that in RPTC: 1) cisplatin-induced mitochondrial dysfunction, decreases in active Na+ transport, and apoptosis are mediated by PKC-alpha and ERK1/2; 2) PKC-alpha and ERK1/2 mediate activation of caspase-3 by acting downstream of cytochrome c release from mitochondria; and 3) ERK1/2 activation by cisplatin occurs through a PKC-alpha-independent pathway.     

Book Review

Salmon, M. V. Practical phase-contrast microscopy. The Microscope, 6, 177-88. 1947.

Evans, Titus C. Radioautographs in which the tissue is mounted directly on the photographic plate. Proc. Soc. Exp. Biol, and Med, 64, 813-15. 1947.

Tolbert, B. M., and Branch, G. E. K. The spectra of the doubly charged positive ions of some p,p'-diaminotriphenylmethane dyes. J. Amer. Chem. Soc, 69, 1083-81. 1947.

Van Wyk, J. J., and Clark, W. M. The luminosity and chromaticity of indicators as a function of pH. J. Amer. Chern. Soc, 69, 1296-1301. 1947.

Hamre, Christopher J. Hematopoiesis in the bone marrow of rats recovering from nutritional anemia. J. Lab. & Clin. Med, 32, 756-76. 1947.

Limarzi, Louis R. Evaluation of bone marrow concentration techniques. A modified method for the simultaneous preparation and staining of blood and bone marrow films. J. lab. & Clin. Med, 32, 782-40. 1947.

Heath, O. V. S. Role of starch in light-induced stomatal movement, and a new reagent for staining stomatal starch. Nature, 159, 647-8. 1947.

Cohen, H. A. A new quick method for staining Treponema pallidum. Acta Med. Orientalia, 6, 99-100. 1947.

Henry, H., and Stacey, M. Histochemistry of the Gram-staining reaction for micro-organisms. Proc. Roy. Soc, Ser. B. Biol. Sci, 133, 891-406. 1946.

Para, M. Silver impregnation of spirochetes in tissue sections. Arch. Path, 42, 649. 1946.

Ruiz, Merino, J. A method of staining capsules. Rev. Sanidad e Hig. Publ (Madrid), 20, 1112. 1946.