Dr. Ron’s Research Review – January 22, 2014

© 2014

This week’s research review focuses on niacinamide and platelets.

Positive

Megakaryocytopoiesis is the process that leads to the production of platelets. This process involves the commitment of multipotent hematopoietic stem cells toward megakaryocyte (MK) progenitors, the proliferation and differentiation of MK progenitors, the polyploidization of MK precursors and the maturation of MK. (Chang, Bluteau et al. 2007)

Nicotinamide (vitamin B3) increases the polyploidisation and proplatelet formation of cultured primary human megakaryocytes. (Giammona, Fuhrken et al. 2006)

Negative

Nicotinamide has been shown to enhance endomitosis in megakaryocytes cultured in vitro, suggesting that it may be beneficial for the production of platelets in culture. However, we found that platelet counts were reduced by about 25% with daily injections of nicotinamide. Although nicotinamide increases polyploidization of megakaryocytes in culture, it does not have translatable effects in vivo. (Konieczna, Panuganti et al. 2013)

Oral nicotinamide reduces serum phosphorus, increases HDL, and induces thrombocytopenia in hemodialysis patients. (Shahbazian, Zafar Mohtashami et al. 2011)

Dr. Ron


Articles

From hematopoietic stem cells to platelets

         (Chang, Bluteau et al. 2007) Download

Megakaryocytopoiesis is the process that leads to the production of platelets. This process involves the commitment of multipotent hematopoietic stem cells toward megakaryocyte (MK) progenitors, the proliferation and differentiation of MK progenitors, the polyploidization of MK precursors and the maturation of MK. Mature MK produce platelets by cytoplasmic fragmentation occurring through a dynamic and regulated process, called proplatelet formation, and consisting of long pseudopodial elongations that break in the blood flow. Recent insights have demonstrated that the MK and erythroid lineages are tightly associated at both the cellular and molecular levels, especially in the transcription factors that regulate their differentiation programs. Megakaryocytopoiesis is regulated by two types of transcription factors, those regulating the differentiation process, such as GATA-1, and those regulating proplatelet formation, such as NF-E2. The humoral factor thrombopoietin (TPO) is the primary regulator of MK differentiation and platelet production through the stimulation of its receptor MPL. Numerous acquired or congenital pathologies of the MK lineage are now explained by molecular abnormalities in the activity of the transcription factors involved in megakaryocytopoiesis, in the Tpo or c-mpl genes, as well as in signaling molecules associated with MPL. The recent development of MPL agonists may provide efficient agents for the treatment of some thrombocytopenias.

Nicotinamide (vitamin B3) increases the polyploidisation and proplatelet formation of cultured primary human megakaryocytes

         (Giammona, Fuhrken et al. 2006) Download

Megakaryocytic (Mk) cell maturation involves polyploidisation, and the number of platelets produced increases with Mk DNA content. Ploidy levels in cultured human MK cells are much lower than those observed in vivo. This study demonstrated that adding the water-soluble vitamin nicotinamide (NIC) to mobilised peripheral blood CD34+ cells cultured with thrombopoietin (Tpo) more than doubled the percentage of high-ploidy (> or = 8N) MK cells. This was observed regardless of donor-dependent differences in Mk differentiation. Furthermore, MK cells in cultures with NIC were larger, had more highly lobated nuclei, reached a maximum DNA content of 64N (vs. 16N with Tpo alone), and exhibited more frequent and more elaborate cytoplasmic extensions. NIC also increased the ploidy of cultured primary murine MK cells and a cell line model (CHRF-288) of Mk differentiation. However, NIC did not alter Mk commitment, apoptosis, or the time at which endomitosis was initiated. Despite the dramatic phenotypic differences observed with NIC addition, gene expression microarray analysis revealed similar overall transcriptional patterns in primary human Mk cultures with or without NIC, indicating that NIC did not disrupt the normal Mk transcriptional program. Elucidating the mechanisms by which NIC increases Mk maturation could lead to advances in the treatment of Mk and platelet disorders.

Mechanistic studies on the effects of nicotinamide on megakaryocytic polyploidization and the roles of NAD+ levels and SIRT inhibition

         (Giammona, Panuganti et al. 2009) Download

OBJECTIVE: Megakaryocytic cells (Mks) undergo endomitosis and become polyploid. Mk ploidy correlates with platelet production. We previously showed that nicotinamide (NIC) greatly increases Mk ploidy in cultures of human mobilized peripheral blood CD34(+) cells. This study aims to examine the generality of NIC effects, NIC's impact on Mk ultrastructure, and potential mechanisms for the increased ploidy. MATERIALS AND METHODS: We used electron microscopy to examine Mk ultrastructure and flow cytometry to evaluate NIC effects on Mk differentiation and ploidy in mobilized peripheral blood CD34(+) cell cultures under diverse megakaryopoietic conditions. Mk ploidy and NAD(H) content were evaluated for NIC and other NAD(+) precursors. We tested additional inhibitors of the sirtuin (or SIRT) 1 and SIRT2 histone/protein deacetylases and, after treatment with NIC, evaluated changes in the acetylation of SIRT1/2 targets. RESULTS: NIC increased ploidy under diverse culture conditions and did not alter Mk ultrastructure; 6.25 mM NIC increased NAD(+) levels fivefold. Quinolinic acid increased NAD(+) similar to that for 1 mM NIC, but yielded a much smaller ploidy increase. Similar increases in Mk ploidy were obtained using NIC or the SIRT1/2 inhibitor cambinol, while the SIRT2 inhibitor AGK2 moderately increased ploidy. SIRT1/2 inhibition in cells treated with NIC was evidenced by increased acetylation of nucleosomes and p53. Greater p53 acetylation with NIC was associated with increased binding of p53 to its consensus DNA binding sequence. CONCLUSION: NIC greatly increases Mk ploidy under a wide range of conditions without altering Mk morphology. Inhibition of SIRT1 and/or SIRT2 is primarily responsible for NIC effects on Mk maturation.

Administration of nicotinamide does not increase platelet levels in mice

         (Konieczna, Panuganti et al. 2013) Download

Elucidating ways to enhance megakaryopoiesis in vivo would have therapeutic applications for thrombocytopenia and transfusion medicine. Nicotinamide has been shown to enhance endomitosis in megakaryocytes cultured in vitro, suggesting that it may be beneficial for the production of platelets in culture. We hypothesized that regular injections of nicotinamide in mice would also increase platelets in vivo. However, we found that platelet counts were reduced by about 25% with daily injections of nicotinamide. Altering the schedule, duration, or nicotinamide dose did not improve platelet production. Consistent with lower platelet levels, nicotinamide also tended to decrease megakaryocyte frequency in sternum and spleen sections, as well as colony formation in vitro by bone marrow progenitor cells. However, there was no effect on the fraction or ploidy of CD41(+) cells harvested from bone marrow. Together, our results suggest that, although nicotinamide increases polyploidization of megakaryocytes in culture, it does not have translatable effects in vivo.

Oral nicotinamide reduces serum phosphorus, increases HDL, and induces thrombocytopenia in hemodialysis patients: a double-blind randomized clinical trial

         (Shahbazian, Zafar Mohtashami et al. 2011) Download

BACKGROUND: Recently, nicotinamide has been suggested as an effective drug for hyperphosphatemia in hemodialysis patients. The authors assessed the efficacy and safety of nicotinamide in these patients with lower doses and longer duration than other studies. METHODS: Forty eight patients with fasting serum phosphorus >5 mg/dl enrolled in this randomized clinical trial study and were randomly assigned to two equal-sized groups of nicotinamide or placebo. The study lasted 8 weeks. In the first four weeks, nicotinamide was administered at 500 mg/day, and in the second four weeks at 1,000 mg/day. Blood samples were tested at baseline, week 4, and week 8. RESULTS: In nicotinamide group, the mean phosphorus level decreased from 5.9 +/- 0.58 mg/dl to 4.77 +/- 1.43 mg/dl in week 4 (P = 0.002) and to 4.66 +/- 1.06 mg/dl in week 8 (P = 0.000). The mean calcium-phosphorus product decreased significantly with the same pattern as phosphorus. High-density lipoprotein level increased from 42.46 +/- 8.01 mg/dl to 55.71 +/- 11.88 mg/dl in week 4 (P = 0.000) and to 65.25 +/- 20.18 mg/dl in week 8 (P = 0.000). Levels of serum calcium, uric acid, SGOT, SGPT, and iPTH didn't change significantly. Compared to baseline, the platelet counts were decreased in both week 4 and week 8. No significant changes were observed in placebo group. CONCLUSIONS: In our patients, nicotinamide effectively decreased phosphorus, increased high-density lipoprotein, and caused thrombocytopenia. Since nicotinamide lowered platelet counts and caused thrombocytopenia in lower doses than other studies in these patients, it is necessary to plan other studies for assessing the safety of the drug especially in different populations.


References

Chang, Y., D. Bluteau, et al. (2007). "From hematopoietic stem cells to platelets." J Thromb Haemost 5 Suppl 1: 318-27. [PMID: 17635743]

Giammona, L. M., P. G. Fuhrken, et al. (2006). "Nicotinamide (vitamin B3) increases the polyploidisation and proplatelet formation of cultured primary human megakaryocytes." Br J Haematol 135(4): 554-66. [PMID: 17054670]

Konieczna, I. M., S. Panuganti, et al. (2013). "Administration of nicotinamide does not increase platelet levels in mice." Blood Cells Mol Dis 50(3): 171-6. [PMID: 23265740]