The Impact of Vitamin D and ATRA on MHC Class II Expression Levels

This presentation investigates the effect of Vitamin D and retinoic acid (ATRA) on Major Histocompatibility Complex (MHC) class II expression, a key component of the adaptive immune system. The study hypothesizes that Vitamin D and ATRA influence HLA-DR expression, impacting autoimmunity. The methods involve harvesting and culturing KG1 cells, fixation, permeabilization for HLA-DR, and analysis using flow cytometry with CD38 live staining. Results indicate that while Vitamin D and ATRA alone have negligible effects on MHC expression, their combined application shows a positive increase in DR and CD38 expression. The conclusion suggests that antigen expression is enhanced when Vitamin D and ATRA are added together. The presentation includes bar charts illustrating median fluorescent intensity for various treatments, providing a visual representation of the experimental findings. Desklib provides access to similar presentations and solved assignments for students.

Effect of Vitamin D on
MHC class II expression
Name of the Student:

Introduction
 Major Histocompatibility complex (MHC) is an integral part of the immune system which
mainly helps in the regulation of the cell mediated adaptive immunity system(Johnson et
al.2016).
 There is a difference between class I and Class II MHCs for the presentation of intracellular
and extracellular antigens respectively (Wieczorek et al.,2017).
 Vitamin D is an essential nutrient which promotes calcium absorption in the gut and diseases
like rickettsia are prevented through sufficient Vitamin D intake.
 According to Andorko et al(2016), HLA-DR molecules carry important immunogenic
domains which induces a humoral immune response.
 Antigen presenting cells (APC) are important for forming complexes with MHCII molecules
leading to antigen exposure and presentation (Finkielsztein2015).

Hypothesis
HLA-DR expression is
affected by Vitamin D
and ATRA which
profoundly affects
auto-immunity.
HLA-DR antigens are
induced by Vitamin D
which further helps in
proper prognosis of
auto immune diseases.
Thus Vitamin D and
ATRA helps in antigen
expression and
processing (Van der
Mejiden et al.2016).

Methods
Harvesting the
cells
Culturing of
KG1 cells
Live staining for
CD38
AML cell line
used
Analysis using
flow cytometry
Fixation and
permiabilizatio
n for HLA-DR
Flow cytometry method

Results
Untreated Isolated With DR
0
20000
40000
60000
80000
100000
120000
140000
160000
180000
Effect of DMSO, RA, Vit D and Vit D, RA both
DMSO
ATRA
VitD
Both
Treatment
Median Fluoroscent Intensity()MFI)

Results
Untreated Isolated CD 38
0
20000
40000
60000
80000
100000
120000
Effect of Vitamin D and ATRA with CD38
DMSO
ATRA
VitD
Both
treatment
Median fluorescent intensity

For checking the effect of the cellular response to stimuli,
staining with antigen was done . Protein expression for
Vitamin D as well as Retinoic acid have also been observed.
CD38 have been used as a control for these experiments. Live
staining procedures have been used the antibody do not bind
epitopes which is on CD38 which can be further modified
through fixation procedures. Hence the samples have been
split.

Discussion and Conclusion
 Vitamin D and Retinoic acid affects the MHC expression in negligible amount.
 Addition of DR has a positive increase upon the addition of Vitamin D and
Retinoic Acid
 Similar increase is seen for CD38 when Vitamin D and Retinoic Acid is added
 From the conclusion it can e understood that the expression of the antigens are
less alone but increases when Vitamin D and Retinoic Acid is added together.

References
 Johnson, D.B., Estrada, M.V., Salgado, R., Sanchez, V., Doxie, D.B., Opalenik, S.R., Vilgelm, A.E., Feld, E.,
Johnson, A.S., Greenplate, A.R. and Sanders, M.E., 2016. Melanoma-specific MHC-II expression represents a
tumour-autonomous phenotype and predicts response to anti-PD-1/PD-L1 therapy. Nature communications, 7,
p.10582.
 Van der Meijden, K., Buskermolen, J., van Essen, H.W., Schuurman, T., Steegenga, W.T., Brouwer-Brolsma, E.M.,
Langenbach, G.E.J., van Ruijven, L.J., den Heijer, M., Lips, P. and Bravenboer, N., 2016. Long-term vitamin D
deficiency in older adult C57BL/6 mice does not affect bone structure, remodeling and mineralization. The
Journal of steroid biochemistry and molecular biology, 164, pp.344-352.
 Wieczorek, M., Abualrous, E. T., Sticht, J., Álvaro-Benito, M., Stolzenberg, S., Noé, F., & Freund, C. (2017).
Major histocompatibility complex (MHC) class I and MHC class II proteins: conformational plasticity in antigen
presentation. Frontiers in immunology, 8, 292.
 Andorko, J. I., Hess, K. L., Pineault, K. G., & Jewell, C. M. (2016). Intrinsic immunogenicity of rapidly-degradable
polymers evolves during degradation. Acta biomaterialia, 32, 24-34.
 Finkielsztein, A., Schlinker, A. C., Zhang, L., Miller, W. M., & Datta, S. K. (2015). Human megakaryocyte
progenitors derived from hematopoietic stem cells of normal individuals are MHC class II-expressing
professional APC that enhance Th17 and Th1/Th17 responses. Immunology letters, 163(1), 84-95.