Influence of Haptoglobin Polymorphism on Stroke in Sickle Cell Disease Patients

This review outlines the current clinical research investigating how the haptoglobin (Hp) genetic polymorphism and stroke occurrence are implicated in sickle cell disease (SCD) pathophysiology. Hp is a blood serum glycoprotein responsible for binding and removing toxic free hemoglobin from the vasculature. The role of Hp in patients with SCD is critical in combating blood toxicity, inflammation, oxidative stress, and even stroke. Ischemic stroke occurs when a blocked vessel decreases oxygen delivery in the blood to cerebral tissue and is commonly associated with SCD. Due to the malformed red blood cells of sickle hemoglobin S, blockage of blood flow is much more prevalent in patients with SCD. This review is the first to evaluate the role of the Hp polymorphism in the incidence of stroke in patients with SCD. Overall, the data compiled in this review suggest that further studies should be conducted to reveal and evaluate potential clinical advancements for gene therapy and Hp infusions.

Acute Kidney Injury in Cardiac Surgery with Cardiopulmonary Bypass

Changes in classification criteria and active introduction of biomarkers of acute kidney injury (KDIGO, 2012) are changing approaches to diagnosis and treatment of postoperative renal dysfunction including cardiac surgery patients operated with cardiopulmonary bypass (CPB). The objective: to compare the detection rate of AKI after surgery with CPB with the use of biomarkers and kidney disease improving global outcomes criteria, as well as to evaluate the cause and localization of structural changes of the nephron.Subjects and Methods. A monocenter observational study among elective cardiac surgery patients (n = 97) was conducted. Inclusion criteria: age over 18 years, duration of surgery (coronary bypass surgery, prosthetic heart valves) from 90 to 180 minutes, no signs of end stage kidney disease. AKI was diagnosed based on changes in serum creatinine and biomarkers (NGAL, IgG, albumin in urine). The studied parameters were recorded 15 minutes after the start and end of anesthesia, as well as 24 and 48 hours after surgery. Retrospectively, the group was divided into three subgroups: 1) patients without AKI after surgery; 2) patients in whom signs of AKI were detected after 24 hours but regressed by the 48th hour; 3) patients in whom AKI persisted during all 48 hours of follow-up.Results. 24 hours after surgery, AKI based on KDIGO criteria was recorded in 56.3% of patients. Using biomarkers, signs of tubular damage (NGAL) at the end of anesthesia were detected in 95.9% of patients; after 24 hours, they were registered in 73.2% of cases. In a subgroup where AKI persisted for more than 24 hours, glomeruli were damaged in addition to tubules which was manifested not only by selective but also by non-selective proteinuria. The duration of CPB, hemodilution (Hb < 90 g/l), the release of free hemoglobin in the blood (> 1.5 mg/l) at low (< 1 g/l) values of haptoglobin were significantly associated with AKI development.Conclusion. The KDIGO criteria do not allow detecting a subclinical form of renal dysfunction which may occur in about 40% of patients after surgery with CPB. AKI can be caused by damage to both the tubular part of the nephron and glomeruli in cases of prolonged CPB with the development of hemolysis, the release of free hemoglobin in the blood, and persisting anemia at the end of the surgery. The NGAL assessment makes it possible to detect subclinical kidney injury in the absence of elevated serum creatinine levels.

Improved Hemolytic Anemia with GBT1118 Is Reno-Protective in Transgenic Sickle Mice

Intravascular hemolysis releases cell-free hemoglobin and heme into the circulation. When haptoglobin and hemopexin are depleted, as observed in sickle cell anemia, cell-free hemoglobin and heme can cause direct oxidative injury, upregulation of inflammatory and immune response pathways, and depletion of nitric oxide. The kidneys are the primary route for clearance of non-scavenged cell-free hemoglobin and heme and are particularly susceptible to these deleterious pathways (PMID 31455889). In patients with sickle cell anemia, increased markers of hemolysis and hemoglobinuria are risk factors for kidney disease and kidney disease progression (PMID 24329963). Voxelotor is a small molecule allosteric modulator that binds and maintains sickle hemoglobin in the oxygenated state, thereby preventing hemoglobin S polymerization and red blood cell sickling. In a phase 3 study, voxelotor improved the degree of hemolysis, as reflected by a rise in hemoglobin concentration and a reduction in indirect bilirubin and reticulocyte percentage (PMID 31199090). The benefits of reducing hemolytic anemia with voxelotor on kidney function are unknown. We investigated whether GBT1118, an analogue of voxelotor with improved pharmacokinetic properties in transgenic sickle mice, would improve biomarkers of kidney damage and kidney function in sickle cell anemia. Transgenic sickle mice (Hb SS; Townes model, Jackson Laboratory) were treated with either GBT1118 (5 male, 5 female) or control (5 male, 5 female) chow from 12 to 24 weeks of age. Urine was collected using metabolic cages for 24 hours at 3 week intervals and blood was collected at baseline, 6 weeks, and 12 weeks of therapy. We compared markers of hemolysis (hemoglobin, reticulocyte percentage), reactive oxygen species (thibarbituric acid reactive substances [TBARS], kidney injury (kidney injury molecule-1 [KIM-1], nephrin), and kidney function (urine albumin and protein, serum blood urea nitrogen [BUN] and creatinine) at the respective time points using ANOVA and adjusting for gender. Mean and standard error values are provided. Consistent with the clinical results observed in the voxelotor phase 3 study, we observed significant improvements in the hemoglobin concentrations and reticulocyte percentages of Hb SS mice treated with GBT1118 versus control (Figure 1A and 1B). The hemoglobin occupancy of GBT1118 after 6 weeks (33 ± 2%) and 12 weeks (28 ± 2%) of therapy were also similar to what was observed in the phase 3 clinical study. An improvement in hemolysis led to reduced hemoglobinuria (6 weeks: treated = 0.07 ± 0.03 vs. control = 0.72 ± 0.2 ng/day, P = 0.004; 12 weeks: treated = 0.04 ± 0.03 vs. control = 1.06 ± 0.32 ng/day, P = 0.006) and less oxidant damage in the kidneys, assessed by urine TBARS concentration, in the treated versus control mice (6 weeks: treated = 31 ± 6 vs. control = 60 ± 10 nmol/day, P = 0.001; 12 weeks: treated = 29 ± 4 vs. control = 71 ± 9 nmol/day, P &lt; 0.001). At 24 weeks of age, markers of glomerular (nephrin: treated = 3.5 ± 0.7 vs. control = 8.5 ± 1.6 ng/day) and tubular injury (KIM-1: treated = 169.6 ± 19.4 vs. control = 348.9 ± 48.2 pg/day) were significantly improved in the treated versus control Hb SS mice (P&lt; 0.001) (Figure 1C). Albuminuria remained stable in the treated Hb SS mice and was relatively close in value to the degree of albuminuria observed in Hb AA mice (Figure 1D). In contrast, albuminuria progressively increased with older age in the control Hb SS mice. A similar pattern was observed for proteinuria (24 week: Hb SS treated = 2.02 ± 0.17 mg/day, Hb SS control = 3.65 ± 0.44 mg/day, Hb AA = 1.65 ± 0.27 mg/day). At 24 weeks of age, the treated Hb SS mice had serum BUN and creatinine values that were similar to what was observed at 12 weeks of age in the Hb SS mice and to the 24-week old Hb AA mice (Figure 1E, 1F). Furthermore, the values for serum BUN and creatinine were significantly lower in the treated versus control Hb SS mice at 24 weeks of age (P ≤ 0.04). In conclusion, we demonstrate that an improvement in hemolysis results in preserved kidney function in transgenic sickle mice. Our findings highlight the clinical importance of hemolytic anemia in the pathophysiology of sickle cell nephropathy. Our results also provide support for developing strategies to mitigate hemolysis in sickle cell anemia in order to provide a targeted approach to improve kidney disease, a devastating complication associated with high morbidity and mortality, in sickle cell anemia. Figure 1 Figure 1. Disclosures Gordeuk: Modus Therapeutics: Consultancy; Novartis: Research Funding; Incyte: Research Funding; Emmaus: Consultancy, Research Funding; Global Blood Therapeutics: Consultancy, Research Funding; CSL Behring: Consultancy. Saraf: Global Blood Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Research Funding.

Haptoglobin 1-1 Isoform Predicts Higher Serum Haptoglobin Concentration and Lower Multiorgan Failure Risk in Sickle Cell Disease

Haptoglobin (HP) is the major scavenger of cell-free hemoglobin in circulation. When haptoglobin is depleted, cell-free hemoglobin can lead to organ damage through direct oxidative injury, upregulation of inflammatory pathways, and depletion of nitric oxide. A common polymorphism in HP gives rise to structurally and functionally distinct phenotypes: HP 1-1, HP 2-1, and HP 2-2. In people of African descent, the HP 1-1 and 2-1 phenotypes have been associated with higher serum HP concentrations compared to the HP 2-2 phenotype (PMID 10807979). In hemolytic diseases, such as sickle cell disease (SCD), the consequence of the HP polymorphism is unclear but could influence cell-free hemoglobin processing and susceptibility to organ damage. We first investigated whether the HP isoforms are associated with HP concentrations at steady-state. Between 10/2009 and 6/2018, we recruited 431 SCD patients into a longitudinal registry at our institution during a routine clinic visit, which we defined as steady-state. Baseline clinical and laboratory data were collected at the time of enrolment. The median age of the cohort was 32 (interquartile range [IQR], 24 - 43) years, 43% were male, 76% were SCD SS or Sβ 0-thalassemia genotype, and 46% were on hydroxyurea at the time of enrolment. Genotyping of the HP polymorphisms by PCR demonstrated the HP 1-1 isoform in 30% (n = 129), the HP 2-1 isoform in 47% (n = 203), and the HP 2-2 isoform in 23% (n = 99) of the SCD cohort. Serum samples were available in 244 of the 431 SCD patients at steady-state. Serum concentrations of HP, measured by ELISA, were higher in those with the HP 1-1 isoform compared to the HP 2-1 or 2-2 isoform (Figure). In a linear regression model, independent variables associated with steady-state serum HP concentration included SCD SS or Sβ 0-thalassemia genotype (β -1.8; 95% CI: -2.5 to -1.0; P &lt; 0.001), LDH (natural log β -0.9; 95% CI: -0.2 to -1.6; P = 0.018), hydroxyurea use (β 0.6; 95% CI: 0.1 to 1.1; P = 0.02), and HP 1-1 isoform (β 0.5, 95% CI: 0 to 1.1; P = 0.049), adjusting for age and sex. Between 9/2020 and 5/2021, we collected an additional 38 serum samples from SCD patients during a hospitalization for vaso-occlusive crisis (VOC) and serum samples from 7 SCD patients during a hospitalization for acute chest syndrome. The HP 1-1 isoform was associated with higher serum HP concentrations during VOC and during acute chest syndrome compared to the HP 2-1 or 2-2 isoforms (Figure). Interestingly, the serum HP trended higher from steady-state to VOC and to acute chest syndrome in SCD patients with the HP 1-1 isoform (β +1.1, P = 0.085) but trended lower in those with the HP 2-1 or 2-2 isoform (β -0.5, P = 0.099). Next we tested whether the HP 1-1 isoform is protective against multi-organ failure during hospitalizations for acute chest syndrome. For the longitudinal analysis, we focused on SCD patients with &gt; 6 months of follow up at our institution (n = 391). A multiorgan failure event was defined as organ dysfunction involving at least 2 of the following organ systems: lung, kidney, liver, or central nervous system (PMID 8109600). With a median follow up of 6.8 (IQR, 3.5 - 8.9) years, we observed an acute chest syndrome episode complicated by multiorgan failure in 14% (53/391) of SCD patients. A significantly lower proportion of SCD patients with the HP 1-1 isoform developed a multiorgan failure event versus those with the 2-1 or 2-2 isoforms (5.7% vs. 17.2%, respectively; P &lt; 0.001). On logistic regression analysis, acute chest syndrome with multiorgan failure was associated with the HP 1-1 isoform (OR 0.3, 95% CI: 0.1 to 0.6; P = 0.002) and baseline LDH (natural log OR 2.7, 95% CI: 1.2 - 6.3; P = 0.02), after adjusting for age, sex, SCD genotype and hydroxyurea use. In conclusion, we demonstrate that the HP 1-1 isoform is independently associated with higher serum HP concentrations at steady-state as well as during hospitalizations for VOC or acute chest syndrome compared to the HP 2-1 or 2-2 isoforms. Furthermore, the HP 1-1 isoform is protective against developing the devastating multiorgan failure syndrome during acute chest syndrome episodes. Our findings highlight the clinical significance of the HP isoforms and cell-free hemoglobin scavenging in SCD. Future studies assessing HP induction and scavenging function based on HP isoform may help elucidate high-risk patients for aggressive measures, such as rapid initiation of exchange transfusion therapy or HP replacement therapy. Figure 1 Figure 1. Disclosures Gordeuk: Modus Therapeutics: Consultancy; Novartis: Research Funding; Incyte: Research Funding; Emmaus: Consultancy, Research Funding; Global Blood Therapeutics: Consultancy, Research Funding; CSL Behring: Consultancy. Saraf: Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Research Funding; Global Blood Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding.

Haptoglobin Gene Polymorphism among Sickle Cell Patients in West Cameroon: Hematological and Clinical Implications

Haptoglobin is a protein involved in protecting the body from the harmful effects of free hemoglobin. The haptoglobin gene exhibits a polymorphism, and the different genotypes do not have the same capacity to combat the free hemoglobin effects. The present study aimed at determining the polymorphic distribution of haptoglobin in sickle cell patients (SCPs) from West Cameroon and their impact on the hematological parameters, as well as clinical manifestations of the disease severity. Haptoglobin genotype of 102 SCPs (SS) and 115 healthy individuals (60 AA and 55 AS) was determined by allele-specific polymerase chain reaction, and the complete blood count was determined using the AutoAnalyser. Results showed that the genotype Hp2-2 was significantly ( p < 0.05) represented in SS patients (54%) than in controls AA and AS (27% and 29%, respectively), while Hp2-1 was mostly found ( p < 0.05) in AS (42%) and AA (38%), against 15% in SS. The allelic distribution in SS patients was Hp2: 0.613, Hp1S: 0.304, and Hp1F: 0.084. In AA and AS controls, the proportions of the Hp1 and Hp2 alleles were similar (around 0.5 each), with 0.282 for Hp1S and 0.218 for Hp1F in AS and 0.283 for Hp1S and 0.258 for Hp1F in AA. The distribution of the haptoglobin genotypes did not reveal any significant difference across hematological parameters and clinical manifestations of disease severity in SCP and controls. SCP with Hp1S-1F genotype presented the highest level of hemoglobin. Although Hp2-2 was more frequent in SS patients, it appeared not to be related to the hematological parameters and to the disease’s severity. Further investigations are necessary to explore the impact of Hp polymorphism such as antioxidant, lipid profile, and functionality of some tissues in SCP in Cameroon.

The In Situ Human Dental Implantome: A First Appraisal

Protein adsorption is the first fundamental interaction between the human body and a foreign surface. The sum of all proteins in this adherent proteinaceous layer comprises the implant proteome. The in situdental implant proteome (implantome) was eluted from four implants of two wettabilities after a 2-min dipping in the humor operationis of maxillar tooth sockets. A mean number of 2056 different polypeptides per implant were identified according to the Xcorr method (Xcorr score ≥ 1.5, n ≥ 2 peptides). In the top 12 proteins comprising ca. 47% of the total abundance, cell-free hemoglobin (26.7%) was the most abundant, followed by fibrinogen (6.4%) and serum albumin (1.8%) with additional 1,800 lower abundance polypeptides, which contained ca. 34 salivary and a similar number of autoimmunogenic polypeptides. Selective enrichment of cell-free hemoglobin on the implant vs. albumin was estimated to 270 fold