Evolved increases in hemoglobin-oxygen affinity and the Bohr effect coincided with the aquatic specialization of penguins

Dive capacities of air-breathing vertebrates are dictated by onboard O₂ stores, suggesting that physiologic specialization of diving birds such as penguins may have involved adaptive changes in convective O₂ transport. It has been hypothesized that increased hemoglobin (Hb)-O₂ affinity improves pulmonary O₂ extraction and enhances the capacity for breath-hold diving. To investigate evolved changes in Hb function associated with the aquatic specialization of penguins, we integrated comparative measurements of whole-blood and purified native Hb with protein engineering experiments based on site-directed mutagenesis. We reconstructed and resurrected ancestral Hb representing the common ancestor of penguins and the more ancient ancestor shared by penguins and their closest nondiving relatives (order Procellariiformes, which includes albatrosses, shearwaters, petrels, and storm petrels). These two ancestors bracket the phylogenetic interval in which penguin-specific changes in Hb function would have evolved. The experiments revealed that penguins evolved a derived increase in Hb-O₂ affinity and a greatly augmented Bohr effect (i.e., reduced Hb-O₂ affinity at low pH). Although an increased Hb-O₂ affinity reduces the gradient for O₂ diffusion from systemic capillaries to metabolizing cells, this can be compensated by a concomitant enhancement of the Bohr effect, thereby promoting O₂ unloading in acidified tissues. We suggest that the evolved increase in Hb-O₂ affinity in combination with the augmented Bohr effect maximizes both O₂ extraction from the lungs and O₂ unloading from the blood, allowing penguins to fully utilize their onboard O₂ stores and maximize underwater foraging time.

In air-breathing vertebrates, diving capacities are dictated by onboard O₂ stores and the efficiency of O₂ use in metabolizing tissues (1). In fully aquatic taxa, selection to prolong breath-hold submergence and underwater foraging time may have promoted adaptive changes in multiple components of the O₂ transport pathway, including oxygenation properties of hemoglobin (Hb). Vertebrate Hb is a tetrameric protein that is responsible for circulatory O₂ transport, loading O₂ in pulmonary capillaries and unloading O₂ in the systemic circulation via quaternary structural shifts between a high-affinity (predominately oxygenated) relaxed (R) state and a low-affinity (predominately deoxygenated) tense (T) state (2). While this mechanism of respiratory gas transport is conserved in all vertebrate Hbs, amino acid variation in the constituent α- and β-type subunits may alter intrinsic O₂ affinity and the responsiveness to changes in temperature, red cell pH, and red cell concentrations of allosteric cofactors (nonheme ligands that modulate Hb-O₂ affinity by preferentially binding and stabilizing the deoxy T conformation) (3, 4).While the quantity of Hb is typically increased in the blood of diving birds and mammals compared with their terrestrial relatives, there is no consensus on whether evolved changes in Hb-O₂ affinity have contributed to enhanced diving capacity (1). It has been hypothesized that increased Hb-O₂ affinity may improve pulmonary O₂ extraction in diving mammals, thereby enhancing diving capacity (5), but more comparative data are needed to assess evidence for an adaptive trend (6, 7). Experimental measurements on whole blood suggest that the emperor penguin (Aptenodytes forsteri) may have a higher blood-O₂ affinity relative to nondiving waterbirds, a finding that has fostered the view that this is a property characterizing penguins as a group (8–10). However, blood-O₂ affinity is a highly plastic trait that is influenced by changes in red cell metabolism and acid-base balance, so measurements on purified Hb under standard assay conditions are needed to assess whether observed species differences in blood-O₂ affinity stem from genetically based changes in the oxygenation properties of Hb. Moreover, even if species differences in Hb-O₂ affinity are genetically based, comparative data from extant taxa do not reveal whether observed differences are attributable to a derived increase in penguins, a derived reduction in their nondiving relatives, or a combination of changes in both directions.To investigate evolved changes in Hb function associated with the aquatic specialization of penguins, we integrated experimental measurements of whole-blood and purified native Hb with evolutionary analyses of globin sequence variation. To characterize the mechanistic basis of evolved changes in Hb function in the stem lineage of penguins, we performed protein engineering experiments on reconstructed and resurrected ancestral Hb representing the common ancestor of penguins and the more ancient ancestor shared by penguins and their closest nondiving relatives (order Procellariiformes, which includes albatrosses, shearwaters, petrels, and storm petrels) (Fig. 1). These two ancestors bracket the phylogenetic interval in which penguin-specific changes in Hb function would have evolved.Open in a separate windowFig. 1.Diagrammatic phylogeny showing the relationship among Sphenisciformes (penguins), Procellariiformes, and Pelecaniformes. Ancestral Hbs were reconstructed for the two indicated nodes: AncSphen and AncPro (the super order that contains Sphenisciformes and Procellariiformes). Divergence times are adapted from Claramunt and Cracraft (56).     

Decreased affinity and number of transferrin receptors on erythroblasts in the anemia of rheumatoid arthritis

In anemia of chronic disease (ACD) in rheumatoid arthritis (RA) a decreased iron uptake and transferrin binding by erythroblasts are postulated to play a pathophysiological role. To examine whether this is related to changes in transferrin receptor expression by erythroblasts, we studied bone marrow from 5 healthy controls, 5 nonanemic RA patients, and 9 RA patients with ACD. Bone marrow mononuclear cells were incubated with increasing concentrations of ¹²⁵I-transferrin and specific binding data were analyzed by the method of Scatchard. The number of transferrin receptors on erythroblasts from RA patients with ACD was significantly lower as compared to nonanemic RA patients (P < .05) and controls (P < .02). The affinity of the transferrin receptor tended to be lower in ACD. These preliminary data may indicate that transferrin receptor expression by erythroblasts is impaired in ACD. Since the rate of erythroid iron uptake is mainly determined by the number of transferrin binding sites, this may explain a decrease in erythroblast iron availability in ACD in RA.     

Changes in Serum Bicarbonate Levels Caused by Acetate‐Containing Bicarbonate‐Buffered Hemodialysis Solution: An Observational Prospective Cohort Study

Fresenius Medical Care's NaturaLyte dialysate has been associated with increased risk of sudden cardiac death by causing metabolic alkalosis from its acetate content based on retrospective data using pre‐dialysis bicarbonate levels only. The study objective was to measure inter/intra‐dialytic changes in serum bicarbonate and degree of alkalosis conferred by varying concentrations of NaturaLyte bicarbonate dialysate. Thirty‐nine hemodialysis patients were divided into four groups based on prescribed bicarbonate dialysate concentrations; Group 1 (N = 9): 30–32 mEq/L, Group 2 (N = 5): 33–34 mEq/L, Group 3 (N = 10): 35–36 mEq/L, Group 4 (N = 15): 37–40 mEq/L. Serial (pre‐dialysis, immediate post‐dialysis, 2 h post‐dialysis, and 68 h post‐dialysis) bicarbonate levels were measured. Mean pre‐dialysis serum bicarbonate levels (representing 44 h post‐dialysis levels) in all four groups were not statistically different. Pre‐dialysis and 68 h post‐dialysis bicarbonate levels in each group were also not significantly different. However, immediate post‐dialysis and 2 h post‐dialysis bicarbonate levels were significantly increased in all four groups proportional to dialysate dose. There was statistically significant inter‐group bicarbonate level difference (P < 0.05) except between the first and second (P = 0.43) and second and third (P = 0.07) groups in the immediate post‐dialysis period. Similar results were obtained for the 2 h post‐dialysis period. High bicarbonate dialysate causes large and rapid fluctuations in serum bicarbonate levels during the intra/inter‐dialytic period, which returns to baseline within 44 to 68 h after dialysis. This refutes the necessity to correct pre‐dialysis acidosis with high bicarbonate dialysate since rapid equilibration is likely to occur and unnecessarily exposes patients to large shifts in their acid base balance.     

Hemolytic anemia provoked by recombinant alpha-interferon

A 30-year-old female with chronic hepatitis C treated with recombinant alpha-interferon developed serious hemolytic anemia after receiving 10 million units a day for 4 weeks. The results of Coombs' tests were negative before and after interferon therapy. The hemolytic anemia gradually improved after the with-drawal of recombinant alpha-interferon and the administration of methyl-prednisolone. The clinical course suggested that the recombinant alpha-interferon had provoked the hemolytic anemia.     

An intermolecular base triple as the basis of ligand specificity and affinity in the guanine- and adenine-sensing riboswitch RNAs

Riboswitches are highly structured RNA elements that control the expression of many bacterial genes by binding directly to small metabolite molecules with high specificity and affinity. In Bacillus subtilis, two classes of riboswitches have been described that discriminate between guanine and adenine despite an extremely high degree of homology both in their primary and secondary structure. We have identified intermolecular base triples between both purine ligands and their respective riboswitch RNAs by NMR spectroscopy. Here, specificity is mediated by the formation of a Watson-Crick base pair between the guanine ligand and a C residue or the adenine ligand and a U residue of the cognate riboswitch RNA, respectively. In addition, a second base-pairing interaction common to both riboswitch purine complexes involves a uridine residue of the RNA and the N3/N9 edge of the purine ligands. This base pairing is mediated by a previously undescribed hydrogen-bonding scheme that contributes to the affinity of the RNA-ligand interaction. The observed intermolecular hydrogen bonds between the purine ligands and the RNA rationalize the previously observed change in specificity upon a C to U mutation in the core of the purine riboswitch RNAs and the differences in the binding affinities for a number of purine analogs.     

New Method for the Approximation of Corrected Calcium Concentrations in Chronic Kidney Disease Patients

The following conventional calcium correction formula (Payne) is broadly applied for serum calcium estimation: corrected total calcium (TCa) (mg/dL) = TCa (mg/dL) + (4 – albumin (g/dL)); however, it is inapplicable to chronic kidney disease (CKD) patients. A total of 2503 venous samples were collected from 942 all‐stage CKD patients, and levels of TCa (mg/dL), ionized calcium ([iCa²⁺] mmol/L), phosphate (mg/dL), albumin (g/dL), and pH, and other clinical parameters were measured. We assumed corrected TCa (the gold standard) to be equal to eight times the iCa²⁺ value (measured corrected TCa). Then, we performed stepwise multiple linear regression analysis by using the clinical parameters and derived a simple formula for corrected TCa approximation. The following formula was devised from multiple linear regression analysis: Approximated  corrected TCa (mg/dL) = TCa + 0.25 × (4 ? albumin) + 4 × (7.4 ? p H) + 0.1 × (6 ? phosphate) + 0.3. Receiver operating characteristic curves analysis illustrated that area under the curve of approximated corrected TCa for detection of measured corrected TCa ≥ 8.4 mg/dL and ≤ 10.4 mg/dL were 0.994 and 0.919, respectively. The intraclass correlation coefficient demonstrated superior agreement using this new formula compared to other formulas (new formula: 0.826, Payne: 0.537, Jain: 0.312, Portale: 0.582, Ferrari: 0.362). In CKD patients, TCa correction should include not only albumin but also pH and phosphate. The approximated corrected TCa from this formula demonstrates superior agreement with the measured corrected TCa in comparison to other formulas.     

A Randomized Controlled Study on the Effects of Acetate‐Free Biofiltration on Organic Anions and Acid‐Base Balance in Hemodialysis Patients

Metabolic acidosis correction is achieved by the transfer of bicarbonate and other buffer anions in dialysis. The aim of this study was to evaluate changes in the main anions of intermediary metabolism on standard hemodiafiltration (HDF) and on acetate‐free biofiltration (AFB). A prospective, in‐center, crossover study was carried out with 22 patients on maintenance dialysis. Patients were randomly assigned to start with 12 successive sessions of standard HDF with bicarbonate (34 mmol/L) and acetate dialysate (3 mmol/L) or 12 successive sessions of AFB without base in the dialysate. Acetate increased significantly during the standard HDF session from 0.078 ± 0.062 mmol/L to 0.156 ± 0.128 mmol/L (P < 0.05) and remained unchanged at 0.044 ± 0.034 mmol and 0.055 ± 0.028 mmol/L in AFB modality. Differences in the acetate levels were observed at two hours (P < 0.005), at the end (P < 0.005) and thirty minutes after the session between HDF and AFB (P < 0.05). There were significantly more patients above the normal range in HDF group than AFB group (68.1% vs 4.5% P < 0.005) postdialysis and 30 minutes later. Serum lactate and pyruvate concentrations decreased during the sessions without differences between modalities. Citrate decreased only in the AFB group (P < 0.05). Acetoacetate and betahydroxybutyrate increased in both modalities, but the highest betahydroxybutyrate values were detected in HDF (P < 0.05). The sum of postdialysis unusual measured organic anions (OA) were higher in HDF compared to AFB (P < 0.05). AFB achieves an optimal control of acid–base equilibrium through a bicarbonate substitution fluid. It also prevents hyperacetatemia and restores internal homeostasis with less production of intermediary metabolites.     

Regional Citrate Anticoagulation for Continuous Renal Replacement Therapy in the Perioperative Care of Liver Transplant Recipients: A Single Center Experience

Kidney injury with concomitant hemodialysis is a common finding in perioperative care of liver transplant patients. The aim of this study was to evaluate disturbances in acid‐base status, electrolyte balance and citrate accumulation during hemodialysis with regional citrate anticoagulation in perioperative care of liver transplant recipients. A retrospective, single center evaluation was conducted of patients with severe liver dysfunction receiving renal replacement therapy in the perioperative care of liver transplantation in a multidisciplinary ICU of a university hospital. Within 5 days of ICU stay, 89 patients undergoing liver transplantation received regional citrate anticoagulation for hemodialysis. During the study period pH (7.39 [7.33/7.43] vs. 7.44 [7.39/7.47], P‐value = 0.014), base excess values (?0.9 [?5.08/2.35] vs. 4.3 [1.93/8.21], P‐value = 0.001) and standard bicarbonate (23.6 [20/26.9] vs. 28.2 [26.2/32.2], P‐value = 0.001) significantly increased, whereas lactate levels (2.6 [1.60/4.45] vs. 1.25 [0.98/1.9], P‐value = 0.071) and Ca_tot/Ca_ion‐ratio decreased or remained below the upper reference. Hypocalcemia appeared mostly within 48 h after dialysis initiation. Although sodium levels increased during the observation, rates of hypernatremia were comparable between hemodialysis days 1 and 5. Hemodialysis using regional citrate anticoagulation remains a challenge in the perioperative care of liver transplant recipients. Major attention must be paid to acid‐base disturbances and citrate accumulation within 48 h after dialysis initiation. Nevertheless, regional citrate anticoagulation in liver dysfunction is a feasible and valuable tool, when limitations and pitfalls are adequately considered.     

Erythropoietin in aplastic anemia

Summary The level of erythropoietin (Ep) was measured in sera and urine from aplastic anemia patients. Increased levels of E_p were demonstrated in sera from all 25 patients studied. An elevated level of E_p was found in the urine of 17 of 23 patients in whom the urine was tested. No correlation between blood hemoglobin and E_p level was observed. A higher serum E_p level was noted in patients with aplastic anemia than in patients with sideropenic anemia of the same severity. To explain the discrepancy, diminished E_p consumption in bone marrow of aplastic anemia patients is discussed.This work was supported by a grant form the Medical Scientific Fund of Serbia     

Normal lysosomal enzyme levels in hypertrophied and failing dog hearts

The activity levels of three lysosomal acid hydrolases, acid phosphatase, acid ribonuclease, and cathepsin, were determined in whole homogenates of right and left ventricular myocardium from normal and chronically stressed dog hearts. Stressed hearts were from four dogs with right ventricular hypertrophy and congestive failure induced by progressive constriction of the main pulmonary artery and from four dogs with acquired right ventricular hypertrophy secondary to pulmonary hypertension resulting from infestation with heartworms (Dirofilaria immitis). Control hearts were from four unoperated and four sham operated normal dogs. No significant differences were found that could be related to myocardial stress.     

Development of alcoholic fatty liver and fibrosis in rhesus monkeys fed a low n-3 fatty acid diet

BACKGROUND: The amount and type of dietary fat seem to be important factors that modulate the development of alcohol-induced liver steatosis and fibrosis. Various alcohol-feeding studies in animals have been used to model some of the symptoms that occur in liver disease in humans. METHODS: Rhesus monkeys (Macaca mulatta) were maintained on a diet that had a very low concentration of alpha-linolenic acid and were given free access to an artificially sweetened 7% ethanol solution. Control and ethanol-consuming animals were maintained on a diet in which the linoleate content was adequate (1.4% of energy); however, alpha-linoleate represented only 0.08% of energy. Liver specimens were obtained, and the fatty acid composition of the liver phospholipids, cholesterol esters, and triglycerides of the two groups were compared at 5 years and histopathology of tissue samples were compared at 3 and 5 years. RESULTS: The mean consumption of ethanol for this group over a 5-year period was 2.4 g.kg.day. As a consequence of the ethanol-dietary treatment, there were significantly lower concentrations of several polyunsaturated fatty acids in the liver phospholipids of the alcohol-treated group, including arachidonic acid and most of the n-3 fatty acids and particularly docosahexaenoic acid, when compared with dietary controls. Liver specimens from animals in the ethanol group at 5 years showed a marked degree of steatosis, both focal and diffuse cellular necrosis, and an increase in the development of fibrosis compared with specimens obtained at 3 years and with those from dietary controls, in which there was no evidence of fibrotic lesions. CONCLUSION: These findings suggest that the advancement of ethanol-induced liver disease in rhesus monkeys may be modulated by the amount and type of dietary essential fatty acids and that a marginal intake of n-3 fatty acids may be a permissive factor in the development of liver disease in primates.     

心-肾贫血综合征的研究进展

慢性肾功能不全的患者常有贫血、心力衰竭,贫血使心力衰竭加重,又使肾功能恶化,心力衰竭控制不好又使肾功能恶化、贫血加重,这三种状态构成心-肾贫血综合征.积极使用促红细胞生成素,纠正贫血,合理使用洋地黄制剂、降压药,适当使用碳酸氢盐透析、超滤纠正心力衰竭、肾衰竭,为临床处理心-肾贫血综合征提供了思路.     

Sideroblastic anemia terminating in myelofibrosis

Two patients with primary acquired sideroblastic anemia who eventually developed myelofibrosis with myeloid metaplasia are reported. Splenectomy was performed in one patient because of increasing blood transfusion requirements, and splenic histology revealed both myeloid metaplasia and ringed sideroblasts. The second patient showed a partial response of the sideroblastic anemia to pyridoxine therapy during the early stages of development of the myelofibrosis. Myelofibrosis with myeloid metaplasia may represent a nonspecific response of the marrow tissue to sideroblastic anemia as to other primary hematologic disorders.     

Functional characteristics of red cells of sickle cell anemia patients with and without α-thalassemia-2

Oxygen equilibrium curves, red cell indices, 2,3-DPG levels, and the percentages of ISC and Hb F were determined for red cell fractions isolated by Dextran 40 density gradient centrifugation from blood of a sickle cell anemia (SS) patient, an SS patient with an additional homozygosity for α-thalassemia-2, and a normal control. In the SS patient, the MCHC and P₅₀ values increased and 2,3-DPG levels decreased with an increase in red cell density; the P₅₀ values were directly related to the MCHC values and inversely related to the 2,3-DPG levels. The bottom fraction contained about 80% ISC, and had the highest MCHC and P₅₀ values and the lowest 2,3-DPG level. In the SS α-thalassemia-2 patient, a decrease in the P₅₀ value corresponded with a decrease in 2,3-DPG levels because no change in MCHC value was observed. The bottom fraction contained about 10% ISC and had the lowest P₅₀ value. These data confirm that both MCHC and 2,3-DPG levels influence the oxygen affinity of SS red cells; an increase in MCHC decreases the oxygen affinity to a great extent while a concomitant decrease in 2,3-DPG level partially abolishes this effect.     

Paravertebral extramedullary hematopoiesis associated with improvement of anemia in congenital dyserythropoietic anemia type II

Thoracic masses resulting from extramedullary hematopoiesis developed in two sisters of Moroccan origin with congenital dyserythropoietic anemia type II (HEMPAS). In one patient, the diagnosis of extramedullary hematopoiesis was confirmed histologically. The appearance of extramedullary foci of hematopoiesis mimicking mediastinal tumors has not been previously described in HEMPAS. These masses result from persistent erythropoietic stimulation associated with chronic hemolytic anemia. In both patients, detection of the asymptomatic masses was preceded by normalization of hemoglobin levels. Thus unexpected correction of a chronic refractory anemia associated with the appearance of mediastinal masses might be the heralding manifestation of an effective extramedullary hemopoiesis.     

Iron-deficiency anemia, non-iron-deficiency anemia and HbA1c among adults in the US

Background: Conditions that affect erythrocyte turnover affect HbA1c concentrations. Although many forms of anemia are associated with lowering of HbA1c, iron deficiency tends to increase HbA1c. We examined the effect of iron and hemoglobin (Hb) status on HbA1c and on the relationship between concentrations of fasting glucose and HbA1c in a national sample of adults in the US. Methods: Cross‐sectional data from 8296 adults aged ≥20 years who participated in NHANES 1999–2002 were used. Results: The prevalence of low Hb (defined as <120 and <118 g/L in women aged 20–69 and ≥70 years, respectively, and <137, <133, and <124 g/L in men aged 20–49, 50–69, and ≥70 years, respectively) was 5.5%. There was a significant positive correlation between Hb concentrations and HbA1c concentrations after adjusting for age, gender, and race or ethnicity, with HbA1c rising from a mean of 5.28% among participants with Hb <100 g/L to 5.72% among participants with Hb ≥170 g/L. The adjusted mean concentrations of HbA1c were 5.56% and 5.46% among participants with and without iron deficiency, respectively (P = 0.095). However, there was no evidence of differences in the relationship between fasting glucose and HbA1c when groups of anemic and non‐anemic individuals with and without iron deficiency were examined individually. Conclusions: Caution should be used when diagnosing diabetes and prediabetes among people with high or low Hb when the HbA1c level is near 6.5% or 5.7%, respectively, as changes in erythrocyte turnover may alter the test result. However, the trend for HbA1c to increase with iron deficiency does not appear to require screening for iron deficiency in ascertaining the reliability of HbA1c in the diagnosis of diabetes and prediabetes in a given individual.     

Acid Secretory Potency and Elimination of the 15-Leucine Gastrin-17 Analogue in Man

The effect on gastric acid secretion and the elimination of a commercially available synthetic human gastrin analogue (15-Leucine Synthetic Human Gastrin I, 15-LSG) were studied in six healthy human subjects. On a molar basis acid secretory efficacy was approximately four times greater than pentagastrin and secretory potency ten times greater. Elimination of 15-LSG was bi-exponential, with half-lives of 5.2 ± 0.8 and 25.9 ± 7.0 min. Metabolic clearance rate was 6.9 ± 0.4 ml/kg-min and volume of distribution 52 ml/kg.     

猪抗人淋巴细胞球蛋白治疗重型再生障碍性贫血的临床研究

目的：探讨一线应用猪抗人淋巴细胞球蛋白（P-ALG）联合环孢素（CsA）治疗重型再生障碍性贫血（SAA）的疗效及不良反应。方法回顾性分析我科应用 P-ALG 联合 CsA 治疗36例 SAA 患者的疗效、并发症及转归,并对患者的年龄、疾病严重程度、发病距应用 P-ALG 的时间等因素与疗效进行相关分析。结果36例患者中死亡4例,白细胞恢复中位时间为30天,脱离红细胞输注的中位时间为55天,脱离血小板输注的中位时间为60天。1年总有效率为86．2％,2年总有效率达91．7％,1年总有效率与性别、年龄、疾病分型、CD8比例无明显相关（P ＞0．05）,病程＜3个月及外周血中性粒细胞（ANC）≥0．2×109／L 患者的有效率高;与国内及欧美国家报道的利用马或兔抗胸腺细胞球蛋白（ATG）治疗 SAA 的有效率相当,而不良反应小,花费仅为马或兔 ATG 治疗的1／3～1／2。结论应用 P-ALG 联合 CsA 治疗重型再生障碍性贫血疗效显著,不良反应轻微。