WHIM syndrome: presumptive diagnosis based on myelokathexis on bone marrow smear

2014 ◽  
Vol 72 (1) ◽  
pp. 111-119 ◽  
Author(s):  
Isabelle Bock ◽  
Franck Dugué ◽  
Elena Loppinet ◽  
Christine Bellanné-Chantelot ◽  
Blandine Bénet
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Smear ◽  
Presumptive Diagnosis ◽  
Whim Syndrome

OXIDATIVE STRESS AND MORPHOLOGICAL ASSESSMENT OF BONE MARROW IN MONOSODIUM GLUTAMATE-TREATED RAT

2018 ◽  
Vol 80 (2) ◽  
Author(s):  
Nor Janna Yahya ◽  
Zariyantey Abd Hamid ◽  
Erni Norfardila Abu Hanipah ◽  
Esther Mathias Ajik ◽  
Nur Afizah Yusoff ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Bone Marrow ◽  
Bone Marrow Cells ◽  
Monosodium Glutamate ◽  
Protein Carbonyl ◽  
Morphological Alteration ◽  
Control Group ◽  
Bone Marrow Smear ◽  
Sprague Dawley ◽  
Oxidative Stress Status

Excess consumption of monosodium glutamate (MSG) was reported to cause oxidative stress on brain, liver and renal and altered haematological parameters. Therefore, this study was aimed to investigate the effect of MSG on oxidative stress status on bone marrow of rats. Male Sprague-Dawley rats (n=24) weighing between 160-200 g were divided randomly into three groups: Control which was given distilled water (1 mg/kg), MSG 60 and MSG 120 which were given 60 mg/kg MSG and 120 mg/kg MSG, respectively. All substances were oral force fed for 28 days consecutively. At the end of the study, bone marrow cells were isolated by flushing technique for measurement of the oxidative stress status and bone marrow smear observation. Results showed that the superoxide dismutase activity and protein carbonyl level were significantly increased in MSG 120 group than to control and MSG 60 groups (p<0.05). Conversely, glutathione level had declined significantly in both MSG groups as compared to control group (p<0.05). The malondialdehyde level was not significantly affected in MSG groups than to control group. Bone marrow smear indicated no evidence of morphological alteration in all groups. In conclusion, MSG at both doses caused oxidative stress on bone marrow after 28 days of exposure.

Incomplete Presentation of WHIM Syndrome: The Diagnostic Role of Dysmorphic Neutrophils in Bone Marrow

2020 ◽  
Vol 42 (7) ◽  
pp. 449-450
Author(s):  
Kousaku Matsubara ◽  
Aya Iwata ◽  
Yu Kawasaki ◽  
Yoshitaka Honda ◽  
Takahiro Yasumi
Keyword(s):  
Bone Marrow ◽  
Whim Syndrome ◽  
Diagnostic Role

Mutant CXCR4 Identified in Neutropenic Patients with Myelokathexis Impairs Survival of Human Myeloid Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3071-3071
Author(s):  
Vahagn Makaryan ◽  
David C. Dale ◽  
Andrew A. Aprikyan
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Cell Survival ◽  
Mutational Analysis ◽  
Bone Marrow Cells ◽  
Myeloid Cells ◽  
Receptor Internalization ◽  
Morphological Examination ◽  
Exon 2 ◽  
Whim Syndrome

Abstract Myelokathexis (WHIM syndrome) is a very rare hematopoietic congenital disorder that is characterized by extremely low level of circulating neutrophils in peripheral blood. It is inherited as an autosomal dominant disease and is diagnosed in early childhood. These patients may have hypogammaglobulinemia and suffer from recurrent infections associated with warts. The hallmark of myelokathexis is a hyperplastic bone marrow and hypersegmented neutrophils with nuclear lobs connected with thin filaments. Myelokathexis is due to a characteristic retention of mature neutrophils in bone marrow, which are not being released to peripheral circulation. We and others reported abnormal cell survival characteristics and impaired bcl-x expression in bone marrow myeloid cells of myelokathexis patients that was partially restored by G-CSF treatment. Recently, it has also been reported that heterozygous truncation mutations in the carboxyterminal domain of the CXCR4 gene, a sole receptor for SDF-1 chemokine, were observed in most, but not all of the families with WHIM syndrome. Subsequently, an impaired receptor internalization and increased chemotaxis towards SDF-1 have been observed in cells expressing truncated CXCR4. Nevertheless, the mechanism of mutant CXCR4 induced myelokathexis remains largely unknown. We performed mutational analysis of the CXCR4 gene in 3 unrelated families with myelokathexis and identified a previously reported R334ter truncation mutation in exon 2 in two of the families. In addition, two silent polymorphisms have been identified in exon 2 of the CXCR4 gene in one of these patients. The third family with afflicted mother and son had a new mutation in the CXCR4 carboxyterminal domain, which resulted in deletion of the last 16 amino acids and subsequent frame shift. None of these mutations were observed in healthy volunteers examined. Since the morphological examination by electron microscopy and flow cytometry analysis of bone marrow cells from some of these patients revealed characteristic apoptotic features, we examined the effect of mutant CXCR4 gene expression on survival of human promyelocytic HL-60 cells. Preliminary data demonstrated that human promyelocytic cells transfected with truncated CXCR4 exhibited impaired cell survival characteristics compared with control HL-60 cells transfected with intact CXCR4. The truncated, but not wild type CXCR4 also increased apoptosis in HL-60 cells induced to differentiate along the granulocytic pathway as determined by flow cytometry of annexin V labeled cells. Thus, these data link together the abnormal survival of proliferating and differentiating myeloid cells in WHIM syndrome with mutant CXCR4 expression. Current studies are focused on elucidation of specific signaling pathways mediating mutant CXCR4-triggered myelokathexis.

Accelerated Apoptosis and Aberrant Retention of Bone Marrow Cells in Myelokathexis Are Two Independent Pathways Triggered by Mutant CXCR4 and Normalized upon Treatment with Specific Inhibitors.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3300-3300
Author(s):  
Vahagn Makaryan ◽  
Oscar Penate ◽  
David Dale ◽  
Andrew Aprikyan
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Mononuclear Cells ◽  
Combined Treatment ◽  
Hematopoietic Cells ◽  
Specific Inhibitor ◽  
Calcium Flux ◽  
Severe Neutropenia ◽  
Pkc Inhibitor ◽  
Whim Syndrome

Abstract Myelokathexis (MK) is a rare congenital hematopoietic disease characterized by hypercellular marrow and severe neutropenia. In some patients there is an association of Warts, Hypogammaglobulinemia, and severe Infections with Myelokathexis (WHIM syndrome). We and others reported accelerated apoptosis of bone marrow myeloid cells in MK evidenced by electron microscopy and flow cytometry studies. The impaired cell survival in MK was associated with reduction in Bcl-X expression at least partially restored by G-CSF treatment. Heterozygous mutations in the CXCR4 gene were reported in most of the families with WHIM syndrome. Interaction of CXCR4, a G-protein coupled chemokine receptor with SDF-1 ligand plays a key role in homing and mobilization of CXCR4-expressing hematopoietic cells. It has been reported that expression of mutant CXCR4 leads to reduced receptor internalization, increased calcium flux and enhanced chemotaxis of transduced CD34+ cells toward SDF1. However, the mechanism of mutant CXCR4-triggered neutropenia in MK remains largely unknown. We examined 11 patients representing 6 unrelated families with MK and found heterozygous CXCR4 mutations in affected but not healthy family members. We identified truncation and deletion mutations including a novel mutation resulting in a deletion of the last 29 amino acids in the cytoplasmic domain of CXCR4. Expression of deletion and truncation mutants in human promyelocytic cells triggered apoptosis similar to that observed in MK patients. Specifically, the rate of apoptotic annexin-positive cells was approximately 2-fold higher in cells transfected with CXCR4 mutants compared with control cells transfected with normal CXCR4. Accelerated apoptosis appeared to stem from a significantly increased dissipation of mitochondrial membrane potential as determined by flow cytometry analysis of DIOC6-labeled cells expressing CXCR4 mutants compared with controls with normal CXCR4 (p<0.02). Similar to enhanced chemotaxis of mononuclear cells observed in MK patients, expression of CXCR4 mutants but not the wild type, resulted in a significant increase in directional motility of cells to SDF-1 (p<0.01). These data indicate that our cellular model appears to closely recapitulate the myelokathexis phenotype. Accelerated apoptosis, but not enhanced chemotaxis induced by mutant CXCR4 was reduced to near-normal level by zVAD-fmk-caspase-specific inhibitor. Interestingly, the mutant CXCR4-triggered increase in chemotaxis to SDF-1 was normalized by treatment with protein kinase C inhibitor. This reduction in mutant CXCR4-mediated increase in chemotaxis in response to the treatment with PKC inhibitor was similar to that observed in response to the treatment with AMD3100. Interestingly, combined treatment with both AMD3100 and PKC inhibitor did not result in a synergistic effect, suggesting that these agents may utilize the same signaling pathway. Of note, neither the treatment with PKC inhibitor not with AMD3100 affected accelerated apoptosis, suggesting that accelerated apoptosis and enhanced chemotaxis are two independent pathways triggered by mutant CXCR4. Importantly, treatment of myelokathexis patients’ blood mononuclear cells with PKC inhibitor restored the abnormal chemotactic properties to near normal levels. Thus, our data suggest that AMD3100 and PKC inhibitor may be effective for treatment of myelokathexis patients with aberrant retention of hematopoietic cells in the bone marrow.

CXCR4 Signals Regulate Neutrophil Release from the Bone Marrow but Not Clearance from the Circulation.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3297-3297
Author(s):  
Kyle J. Eash ◽  
Jacquelyn M. Means ◽  
Jill R. Woloszynek ◽  
Fulu Liu ◽  
Daniel C. Link
Keyword(s):  
Bone Marrow ◽  
Half Life ◽  
Microbial Pathogens ◽  
Minor Role ◽  
Wild Type ◽  
Cxcr4 Signaling ◽  
Whim Syndrome ◽  
Chemokine Cxcl12 ◽  
Post Infusion ◽  
A Minor

Abstract The number of circulating neutrophils is tightly regulated in order to effectively protect against microbial pathogens while minimizing damage to host tissue. Homeostatic control of neutrophils in the blood is achieved through a balance of neutrophil production, release from the bone marrow, and clearance from the circulation. Accumulating evidence suggests that signaling by the chemokine CXCL12, through its major receptor CXCR4, may play a key role in controlling neutrophil homeostasis. Indeed, gain-of-function mutations of CXCR4 are responsible for most cases of WHIM syndrome, a syndrome that features impaired neutrophil release from the bone marrow. Conversely, we previously reported that mice carrying a myeloid-specific deletion of CXCR4 (CXCR4f/−LysM+/Cre mice) display constitutive neutrophil release. Moreover, we provided data suggesting that neutrophil mobilization by G-CSF or Groβ are dependent on CXCR4 signaling, as neutrophil mobilization by these agents was absent in CXCR4f/−LysM+/Cre mice. These data firmly establish CXCR4 signaling as a key regulator of neutrophil release from the bone marrow under basal and stress conditions. Though controversial, there also is evidence that CXCR4 may play a role in neutrophil clearance from the blood by selectively trapping and removing aged neutrophils in the bone marrow. In this study, we examine the role of CXCR4 in neutrophil clearance using CXCR4f/−LysM+/Cre mice. Strain-matched wild type or CXCR4f/−LysM+/Cre mice were treated with a single injection of BrdU to label newly synthesized neutrophils. A similar percentage of myeloid cells in the bone marrow were labeled in wild type and CXCR4f/−LysM+/Cre mice, suggesting that the loss of CXCR4 does not affect granulocytic cell proliferation. Consistent with its role in regulating neutrophil release, the transit time for labeled neutrophils to appear in the circulation was significantly reduced in CXCR4f/−LysM+/Cre mice (45 hours) compared with wild type mice (72 hours). The half-life (t1/2 ) of neutrophils in the blood was calculated using the formula N=N0e−λt where N0 = the peak number of labeled cells, N = the number of cells at time t and λ = the decay constant. Surprisingly, no difference in the circulating neutrophil half-life was observed in CXCR4f/−LysM+/Cre mice compared to wild type mice (18.3 ± 13.6 hours vs.12.7 ± 9.5 hours respectively, P=0.43). We next performed adoptive transfer experiments to determine the site of neutrophil clearance. Specifically, an equivalent number of bone marrow neutrophils from wild type or CXCR4f/−LysM+/Cre mice were injected intravenously into recipient mice. Donor neutrophils were identified based on differential Ly5 gene expression. By 3 hours post-infusion, the majority of donor neutrophils were cleared from the blood. Compared to wild type neutrophils, CXCR4−/− neutrophils showed reduced homing to the bone marrow [number of donor neutrophils per femur: 6.7 ± 0.3 x 104 (wild type) compared to 2.6 ± 0.8 x 104 (CXCR4−/−); P <0.05]. Conversely, an increased number of CXCR4−/− neutrophils were present in the spleen. These data confirm that CXCR4 expression on neutrophils plays a role in the homing of neutrophils back to the bone marrow. However, neutrophil removal in the bone marrow appears to play only a minor role in neutrophil clearance from the blood, as neutrophil half-life was not significantly affected by the loss of CXCR4.

Somatic Activating Mutations In CXCR4 Are Common In Patients With Waldenstrom’s Macroglobulinemia, and Their Expression In WM Cells Promotes Resistance To Ibrutinib

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4424-4424 ◽  
Author(s):  
Yang Cao ◽  
Zachary Hunter ◽  
Xia Liu ◽  
Lian Xu ◽  
Guang Yang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Proliferation ◽  
Somatic Mutation ◽  
Sanger Sequencing ◽  
Somatic Mutations ◽  
Akt Signaling ◽  
Wild Type ◽  
Control Vector ◽  
Whim Syndrome ◽  
Frame Shift

Background Waldenstrom's macroglobulinemia (WM) is an indolent non-Hodgkin's lymphoma characterized by the accumulation of IgM secreting lymphoplasmacytic cells in the bone marrow. CXCR4 is a chemokine receptor that promotes the survival, migration, and adhesion to the bone marrow stroma of WM lymphoplasmacytic cells (LPC) through interactions with its ligand CXCL12. Through whole genome sequencing, we identified somatic mutations in CXCR4 that affected 1/3 of WM patients. These mutations were identical or functionally similar to those associated with Warts, Hypogammaglobulinemia, Infection, and Myelokathexis (WHIM) syndrome (Hunter et al, ASCO 2012), a rare autosomal dominant genetic disorder that is caused by frame shift or nonsense mutations in the carboxyl-terminal cytoplasmic tail of CXCR4. In WHIM syndrome, loss of the c-terminal tail of CXCR4 impairs receptor internalization, thereby prolonging G-protein and β-arrestin signaling (Lugane et al, Blood 2008). Ibrutinib induces WM cell death, and is highly active in WM (Treon et al, ICML-12, 2013). Since the target of ibrutinib (BTK) is a known downstream target of CXCR4, we sought to clarify if ibrutinib activity in WM LPCs was modulated by WHIM-like mutations in CXCR4. Methods We first sought to confirm the frequency of WHIM-like mutations in 87 untreated WM patients by Sanger sequencing. The most common CXCR4 somatic mutation identified (S338X) in these studies was then cloned by PCR from CD19+ LPCs from a WM patient with this somatic mutation. Wild type (WT) and S338X CXCR4 cDNAs were subcloned into plenti-IRES-GFP vector, and transduced using an optimized lentiviral based strategy into BCWM.1 WM cells. Five days after transduction, GFP positive cells were sorted and used for functional studies. Surface expression of CXCR4 was determined by flow cytometeric analysis using a PE-conjugated anti-CXCR4 monoclonal antibody. The expression of phosphorylated BTK, AKT, and ERK1/2 was determined by western blot analysis. Cell proliferation was measured with alamar blue. Results Sanger sequencing identified nonsense or frame shift mutations (WHIM-like) in the c-terminal tail of CXCR4 in 28 of 87 (32%) patients, the most common of which was a non-sense mutation (S338X) that was present in 12 patients. BCWM.1 cells were then transduced with control vector, CXCR4 wild type or CXCR4 S338X mutant expressing vectors. Expression was confirmed by cDNA Sanger sequencing. Stably transduced cells exposed to ibrutinib (0.5uM or 1uM) showed significantly reduced cell proliferation (p<0.005). Ibrutinib treated control vector and CXCR4 wild-type transduced cells showed suppressed tumor cell growth even in the presence of the CXCR4 ligand CXCL12 (20 nM), whereas cells transduced with CXCR4 S338X WHIM-like mutation demonstrated resistance to ibrutinib growth effect (p<0.005). In turn, this rescue could be blocked by treatment with 30uM of the CXCR4 specific inhibitor AMD3100 confirming that this effect was mediated through CXCR4 (p<0.005) (Figure 1). Phosphorylated BTK, ERK1/2 and AKT signaling increased following CXCL12 stimulation in all transduced cells, while ibrutinib inhibited their activation in control vector and CXCR4 wild-type, but not CXCR4 S338X mutant cells. CXCR4 triggered signaling by CXCL12 in these experiments was confirmed by pre-treatment with AMD3100. Conclusions By Sanger sequencing, WHIM-like CXCR4 somatic mutations are observed in 1/3 of untreated WM patients. WHIM-like CXCR4 mutations are associated with resistance to ibrutinib mediated ERK1/2 and AKT signaling, as well as growth suppression in the presence of the CXCR4 ligand, CXCL12, in WM cells. These studies may have important implications for CXCR4 modulation in the treatment of WM, as well as potential use of CXCR4 mutations in predicting outcome for patients undergoing ibrutinib therapy. Disclosures: No relevant conflicts of interest to declare.

Acquired Whim Syndrome: A Possible New Side Effect Of Dasatinib!

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4933-4933 ◽  
Author(s):  
Satya Prakash Yadav ◽  
Mohammed Ramzan ◽  
Yogi Raj Chopra ◽  
Nivedita Dhingra ◽  
Anupam Sachdeva
Keyword(s):  
Bone Marrow ◽  
Maintenance Therapy ◽  
Myeloid Leukaemia ◽  
Immunoglobulin G ◽  
Side Effect ◽  
Serum Immunoglobulin ◽  
Molecular Remission ◽  
Whim Syndrome ◽  
Cxcr4 Receptor ◽  
Tract Infections

Abstract Introduction WHIM (Warts, Hypogammaglobulinemia, Infections and Myelokathexis) syndrome is a hereditary disorder caused by activating mutation in CXCR4 receptor. CXCR4 receptor is expressed in many cells including myeloid and lymphoid cells. Dasatanib is aTKI inhibitor and very effective drug in treating Ph+ve acute lymphoblastic leukaemia (Ph+ALL). Dasatinib has many off target effects which are not completely known as yet. Dasatinib effect on CXCR4 is not well described. In Chronic myeloid leukaemia it causes reacquisition CXCR4 activation leading to hiding of myeloid leukaemia cells inside the bone marrow. However some work has been done on effects of Dasatinib on mature neutrophil indicates adhesion defect and inhibition of pro-inflammatory effect. Here we describe two children who developed acquired WHIM syndrome like features while on Dasatinib therapy. Methods Two girls aged 9 and 11 year old with Ph+ALL were treated initially as per BFM95 protocol and Imatinib 375 mg/m2 daily. Both went in remission and later achieved complete molecular remission. However during maintenance therapy first patient at 19 months from diagnosis and second at 12 months from diagnosis started having rising value of quantitative BCR-ABL values. As both children had no siblings and no matched unrelated donor was available so in both Imatinib was stopped and after taking informed consent of parents Dasatinib was started at a dose of 100/mg/m2/day in two divided doses and maintenance therapy of oral 6-Mercaptopurine (6-MP) and Methotrexate (MTX) was continued. They both achieved reduction in BCR-ABL quantitative PCR after 3 months and molecular remission at 6 and 9 months respectively after starting Dasatinib. Results During therapy with dasatinib and oral 6-MP and MTX both children started having recurrent respiratory tract infections with neutropenia and warts appeared on face and body. Serum immunoglobulin G levels were lower than 200 mg/dl in both. We felt that these symptoms resembled WHIM syndrome and possibly could be due to Dasatinib. So we gave Intravenous immunoglobulin to both the children and IV antibiotics and stopped both Dasatinib and chemotherapy for a week. Neutropenia recovered within 72 hr after stopping of dasatinib. However, we did not perform bone marrow examination in both the cases to prove myelokathexis. Both children did well when Dasatinib was restarted at a lower dose of 50 mg/m2/day once daily and oral 6-MP and MTX. Warts disappeared over next 4-6 months and did not have any further episodes of fever or febrile neutropenia needing IV antibiotics. Their repeat serum immunoglobulin G levels stayed normal over next 6-12 months. Conclusion Both our children on dasatinib developed WHIM syndrome like features which resolved on reducing dose of Dasatinib. Dasatinib possibly mediates this side effect through CXCR4 receptor or downstream pathway. More data are needed to prove this association. Disclosures: No relevant conflicts of interest to declare.

scholarly journals WHIM syndrome myelokathexis reproduced in the NOD/SCID mouse xenotransplant model engrafted with healthy human stem cells transduced with C-terminus–truncated CXCR4

Blood ◽  
2006 ◽  
Vol 109 (1) ◽  
pp. 78-84 ◽  
Author(s):  
Toshinao Kawai ◽  
Uimook Choi ◽  
Lanise Cardwell ◽  
Suk See DeRavin ◽  
Nora Naumann ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Transgene Expression ◽  
Scid Mouse ◽  
Severe Combined Immunodeficiency ◽  
Myeloid Cells ◽  
Xenograft Model ◽  
Healthy Human ◽  
Whim Syndrome ◽  
Mouse Xenograft Model

AbstractWHIM(warts, hypogammaglobulinemia, recurrent bacterial infection, and myelokathexis) syndrome is a rare immunodeficiency caused in many cases by autosomal dominant C-terminal truncation mutations in the chemokine receptor CXCR4. A prominent and unexplained feature of WHIM is myelokathexis (hypercellularity with apoptosis of mature myeloid cells in bone marrow and neutropenia). We transduced healthy human CD34+ peripheral blood–mobilized stem cells (PBSCs) with retrovirus vector encoding wild-type (wt) CXCR4 or WHIM-type mutated CXCR4 and studied these cells ex vivo in culture and after engraftment in a nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mouse xenograft model. Neither wt CXCR4 nor mutated CXCR4 transgene expression itself enhanced apoptosis of neutrophils arising in transduced PBSC cultures even with stimulation by a CXCR4 agonist, stromal cell–derived factor-1 (SDF-1 [CXCL12]). Excess wt CXCR4 expression by transduced human PBSCs enhanced marrow engraftment, but did not affect bone marrow (BM) apoptosis or the release of transduced leukocytes into PB. However, mutated CXCR4 transgene expression further enhanced BM engraftment, but was associated with a significant increase in apoptosis of transduced cells in BM and reduced release of transduced leukocytes into PB. We conclude that increased apoptosis of mature myeloid cells in WHIM is secondary to a failure of marrow release and progression to normal myeloid cell senescence, and not a direct effect of activation of mutated CXCR4.

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