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Type of in vitro cultivation influences cytoadhesion, knob structure, protein localization and transcriptome profile of Plasmodium falciparum.

Tilly AK, Thiede J, Metwally N, Lubiana P, Bachmann A, Roeder T, Rockliffe N, Lorenzen S, Tannich E, Gutsmann T, Bruchhaus I - Sci Rep (2015)

Bottom Line: In addition, there was a greater reduction in the cytoadhesion of IEs to various endothelial receptors in the presence of AlbuMAX than in the presence of human serum.Surprisingly, cytoadhesion did not correlate with the presence or absence of knobs.Greater numbers of the variant surface antigen families RIFIN, STEVOR, and PfMC-2TM were found at the IE membrane when cultivated in the presence of AlbuMAX.

View Article: PubMed Central - PubMed

Affiliation: Bernhard Nocht Institute for Tropical Medicine, Bernhard-Nocht-Str. 74, 20359 Hamburg, Germany.

ABSTRACT
In vitro cultivation of Plasmodium falciparum is critical for studying the biology of this parasite. However, it is likely that different in vitro cultivation conditions influence various aspects of the parasite's life cycle. In the present study two P. falciparum isolates were cultivated using the two most common methods, in which AlbuMAX or human serum as additives are used, and the results were compared. The type of cultivation influenced the knob structure of P. falciparum-infected erythrocytes (IEs). IEs cultivated with AlbuMAX had fewer knobs than those cultivated with human serum. Furthermore, knob size varied between isolates and is also depended on the culture medium. In addition, there was a greater reduction in the cytoadhesion of IEs to various endothelial receptors in the presence of AlbuMAX than in the presence of human serum. Surprisingly, cytoadhesion did not correlate with the presence or absence of knobs. Greater numbers of the variant surface antigen families RIFIN, STEVOR, and PfMC-2TM were found at the IE membrane when cultivated in the presence of AlbuMAX. Moreover, the type of cultivation had a marked influence on the transcriptome profile. Compared with cultivation with human serum, cultivation with AlbuMAX increased the expression of approximately 500-870 genes.

No MeSH data available.


Related in: MedlinePlus

Atomic force microscopy of P. falciparum IEs.Isolates 3D7 and FCR3 were cultivated in the presence of AlbuMAX or human serum (K−). In addition, the isolates were enriched for the presence of knobs by gelatin sedimentation (K+). In each case, highly synchronized trophozoite-stage parasites (28–32 hpi) were examined. A. Scanning measurements from atomic force micrographs of 3D7 and FCR3 IEs, as well as uninfected erythrocytes (control). B. Surface area of the knobs on 3D7 and FCR3 IEs enriched for the presence of knobs. (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001).
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f4: Atomic force microscopy of P. falciparum IEs.Isolates 3D7 and FCR3 were cultivated in the presence of AlbuMAX or human serum (K−). In addition, the isolates were enriched for the presence of knobs by gelatin sedimentation (K+). In each case, highly synchronized trophozoite-stage parasites (28–32 hpi) were examined. A. Scanning measurements from atomic force micrographs of 3D7 and FCR3 IEs, as well as uninfected erythrocytes (control). B. Surface area of the knobs on 3D7 and FCR3 IEs enriched for the presence of knobs. (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001).

Mentions: We next compared the physical appearance of knob-containing IEs in the presence of AlbuMAX or human serum using atomic force microscopy, which allows very high resolution imaging (Fig. 1, Fig. 4A). Although the knob density/μm for isolate 3D7 was lower in the presence of AlbuMAX than in the presence of human serum, there were no differences in the size measured as surface area of the knobs (Fig. 4B). However, when FCR3 was cultivated in the presence of AlbuMAX, the knobs varied greatly in terms of size. The smallest knobs covered a surface area of approximately 3,300 nm2, whereas the largest covered a surface area between 20,000 and 36,000 nm2. The size of the knobs on IEs cultivated in the presence of human serum was more uniform; in this case the knobs covered a surface area between 900 and 14,000 nm2, the average being about 8,000 nm2. Therefore, cultivating FCR3 IEs in the presence of human serum resulted in the formation of smaller, more uniform knobs when compared with cultivation in the presence of AlbuMAX (Fig. 4B, Table 1). The knobs on erythrocytes infected with isolate FCR3 were larger than those on cells infected with isolate 3D7 when cultivated in AlbuMAX. By contrast, the knobs of FCR3 IEs were smaller than those of 3D7 IEs when cultivated in human serum (Fig. 4B). When the height and width of the knobs were analyzed separately, it was clear that the knobs of 3D7 IEs cultivated with AlbuMAX were wider (13 ± 9 nm) than those of FCR3 IEs; however, those of FCR3 IEs were higher (115 ± 40 nm) (Tables 1 and 2). The dried samples were imaged in air. Thus, the measured heights appear lower as they appear in fixed EM sections. In summary, sizes of knobs differed between the two isolates 3D7 and FCR3. While for FCR3 the type of cultivation has a strong influence on knob sizes, the influence of cultivation on knob sizes in 3D7 was not significant.


Type of in vitro cultivation influences cytoadhesion, knob structure, protein localization and transcriptome profile of Plasmodium falciparum.

Tilly AK, Thiede J, Metwally N, Lubiana P, Bachmann A, Roeder T, Rockliffe N, Lorenzen S, Tannich E, Gutsmann T, Bruchhaus I - Sci Rep (2015)

Atomic force microscopy of P. falciparum IEs.Isolates 3D7 and FCR3 were cultivated in the presence of AlbuMAX or human serum (K−). In addition, the isolates were enriched for the presence of knobs by gelatin sedimentation (K+). In each case, highly synchronized trophozoite-stage parasites (28–32 hpi) were examined. A. Scanning measurements from atomic force micrographs of 3D7 and FCR3 IEs, as well as uninfected erythrocytes (control). B. Surface area of the knobs on 3D7 and FCR3 IEs enriched for the presence of knobs. (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4645185&req=5

f4: Atomic force microscopy of P. falciparum IEs.Isolates 3D7 and FCR3 were cultivated in the presence of AlbuMAX or human serum (K−). In addition, the isolates were enriched for the presence of knobs by gelatin sedimentation (K+). In each case, highly synchronized trophozoite-stage parasites (28–32 hpi) were examined. A. Scanning measurements from atomic force micrographs of 3D7 and FCR3 IEs, as well as uninfected erythrocytes (control). B. Surface area of the knobs on 3D7 and FCR3 IEs enriched for the presence of knobs. (*p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001).
Mentions: We next compared the physical appearance of knob-containing IEs in the presence of AlbuMAX or human serum using atomic force microscopy, which allows very high resolution imaging (Fig. 1, Fig. 4A). Although the knob density/μm for isolate 3D7 was lower in the presence of AlbuMAX than in the presence of human serum, there were no differences in the size measured as surface area of the knobs (Fig. 4B). However, when FCR3 was cultivated in the presence of AlbuMAX, the knobs varied greatly in terms of size. The smallest knobs covered a surface area of approximately 3,300 nm2, whereas the largest covered a surface area between 20,000 and 36,000 nm2. The size of the knobs on IEs cultivated in the presence of human serum was more uniform; in this case the knobs covered a surface area between 900 and 14,000 nm2, the average being about 8,000 nm2. Therefore, cultivating FCR3 IEs in the presence of human serum resulted in the formation of smaller, more uniform knobs when compared with cultivation in the presence of AlbuMAX (Fig. 4B, Table 1). The knobs on erythrocytes infected with isolate FCR3 were larger than those on cells infected with isolate 3D7 when cultivated in AlbuMAX. By contrast, the knobs of FCR3 IEs were smaller than those of 3D7 IEs when cultivated in human serum (Fig. 4B). When the height and width of the knobs were analyzed separately, it was clear that the knobs of 3D7 IEs cultivated with AlbuMAX were wider (13 ± 9 nm) than those of FCR3 IEs; however, those of FCR3 IEs were higher (115 ± 40 nm) (Tables 1 and 2). The dried samples were imaged in air. Thus, the measured heights appear lower as they appear in fixed EM sections. In summary, sizes of knobs differed between the two isolates 3D7 and FCR3. While for FCR3 the type of cultivation has a strong influence on knob sizes, the influence of cultivation on knob sizes in 3D7 was not significant.

Bottom Line: In addition, there was a greater reduction in the cytoadhesion of IEs to various endothelial receptors in the presence of AlbuMAX than in the presence of human serum.Surprisingly, cytoadhesion did not correlate with the presence or absence of knobs.Greater numbers of the variant surface antigen families RIFIN, STEVOR, and PfMC-2TM were found at the IE membrane when cultivated in the presence of AlbuMAX.

View Article: PubMed Central - PubMed

Affiliation: Bernhard Nocht Institute for Tropical Medicine, Bernhard-Nocht-Str. 74, 20359 Hamburg, Germany.

ABSTRACT
In vitro cultivation of Plasmodium falciparum is critical for studying the biology of this parasite. However, it is likely that different in vitro cultivation conditions influence various aspects of the parasite's life cycle. In the present study two P. falciparum isolates were cultivated using the two most common methods, in which AlbuMAX or human serum as additives are used, and the results were compared. The type of cultivation influenced the knob structure of P. falciparum-infected erythrocytes (IEs). IEs cultivated with AlbuMAX had fewer knobs than those cultivated with human serum. Furthermore, knob size varied between isolates and is also depended on the culture medium. In addition, there was a greater reduction in the cytoadhesion of IEs to various endothelial receptors in the presence of AlbuMAX than in the presence of human serum. Surprisingly, cytoadhesion did not correlate with the presence or absence of knobs. Greater numbers of the variant surface antigen families RIFIN, STEVOR, and PfMC-2TM were found at the IE membrane when cultivated in the presence of AlbuMAX. Moreover, the type of cultivation had a marked influence on the transcriptome profile. Compared with cultivation with human serum, cultivation with AlbuMAX increased the expression of approximately 500-870 genes.

No MeSH data available.


Related in: MedlinePlus