Oxygen metabolism of mammalian spermatozoa. Generation of hydrogen peroxide by rabbit epididymal spermatozoa

Rabbit spermatozoa from the cauda epididymis produced 0.7–0.8nmol of H₂O₂/min per 10⁸ cells at cell concentrations below 10⁷ cells/ml with linear dependence on cell concentration. Above 2 × 10⁷ cells/ml, the rate again became linear with cell concentration but decreased to 0.1–0.2nmol/min per 10⁸ cells. Spermatozoa treated with amphotericin B, which makes the plasma membrane highly permeable to low-molecular-weight compounds, showed a similar dependence of H₂O₂ production rate on cell concentration; below 10⁷ cells/ml the rate was 0.3–0.4nmol/min per 10⁸ cells; above 2 × 10⁷ cells/ml, the rate was 0.1–0.2nmol/min per 10⁸ cells. Hypo-osmotically treated rabbit epididymal spermatozoa, a preparation useful for studying mitochondrial function in sperm [Keyhani & Storey (1973) Biochim. Biophys. Acta 305, 557–565] produced 0.1–0.2nmol/min per 10⁸ cells in the absence of added substrates. The dependence of rate on cell concentration was linear from 10⁷ to 2.2 × 10⁸ cells/ml. This endogenous rate was unaffected by rotenone, but stimulated 4-fold by antimycin A. Addition of the mitochondrial substrates lactate plus malate increased the rate of H₂O₂ production to 0.3nmol/min per 10⁸ cells. The decreased rate of H₂O₂ production observed with intact sperm at high cell concentrations is attributed to reaction of H₂O₂ with the cells, possibly with the plasma membrane, which is lost after hypo-osmotic treatment. Rabbit spermatozoa have glutathione peroxidase and glutathione reductase activities, but these seem to play little role in removal of H₂O₂ generated. The rate at low cell concentration is taken to be the unperturbed rate. The sources of H₂O₂ production in rabbit spermatozoa have been tentatively resolved into a low-molecular-weight component, lost after amphotericin treatment, a mitochondrial component and a rotenone-insensitive component that has not been identified.     

CHANGE IN BLOOD GELSOLIN CONCENTRATION IN RESPONSE TO PHYSICAL EXERCISE

Plasma gelsolin (pGSN) produced by muscle is an abundant protein of extracellular fluids capable of severing actin filaments and eliminating actin from the circulation. Additionally, pGSN modulates the cellular effects of some bioactive lipids. In this study we test the hypothesis that hormonal and metabolic adaptations to exercise are associated with changes in gelsolin concentration in blood. Plasma samples were collected from twenty healthy males recruited from untrained (UT, n=10) and endurance trained (ET, n=10) groups that performed 30-60 minutes of exercise on a cycloergometer at a workload corresponding to 70% of VO₂max. Gelsolin concentration was determined by quantitative Western blot analysis with an anti-human gelsolin antibody. The gelsolin concentration in UT and ET subjects before starting exercise ranged from 104 to 330 and 163 to 337 µg · ml^-1 respectively. After 30 minutes of exercise we observed a significant decrease of plasma gelsolin in the UT group (p<0.05) while the gelsolin concentration in the ET group rose on average from 244 to 271 µg · ml^-1. However, this increase did not reach statistical significance. Endurance training might increase the ability of muscle tissue to express plasma gelsolin as part of an adaptive mechanism.     

STUDIES ON CELL METABOLISM AND CELL DIVISION : III. OXYGEN CONSUMPTION AND CELL DIVISION OF FERTILIZED SEA URCHIN EGGS IN THE PRESENCE OF RESPIRATORY INHIBITORS

1. The effects of a number of respiratory inhibiting agents on the cell division of fertilized eggs of Arbacia punctulata have been determined. For eggs initially exposed to the reagents at 30 minutes after fertilization at 20°C., the levels of oxygen consumption prevailing in the minimum concentrations of reagents which produced complete cleavage block were (as percentages of the control): In 0.4 per cent O₂-99.6 per cent N₂, 32; in 0.7 per cent O₂-99.3 per cent CO, 32; in 1.6 x 10^–4 M potassium cyanide, 34; in 1 x 10^–3 M phenylurethane, 70; in 4 x 10^–3 M 5-isoamyl-5-ethyl barbituric acid, 20; in 3 x 10^–4 M iodoacetic acid, 53. 2. The carbon monoxide inhibition of oxygen consumption and cell division was reversed by light. The percentage inhibition of oxygen consumption by carbon monoxide in the dark is described by the usual mass action equation with K, the inhibition constant, equal to approximately 60, as compared to values of 5 to 10 for yeast and muscle. In 20 per cent O₂-80 per cent CO in the dark there was a slight stimulation of oxygen consumption, averaging 20 per cent. 3. Spectroscopic examination of fertilized and unfertilized Arbacia eggs reduced by hydrosulfite revealed no cytochrome bands. The thickness and density of the egg suspension was such as to indicate that, if cytochrome is present at all, the amount in Arbacia eggs is extremely small as compared to that in other tissues having a comparable rate of oxygen consumption. 4. Three reagents poisoning copper catalyses, potassium dithio-oxalate (10^–2 M), diphenylthiocarbazone (10^–4 M), and isonitrosoacetophenone (2 x 10^–3 M) produced no inhibition of division of fertilized Arbacia eggs. 5. These results indicate that the respiratory processes required to support division in the Arbacia egg may perhaps differ in certain essential steps from the principal respiratory processes in yeast and muscle.     

THE COMBINATION OF CERTAIN PROTEINS WITH HYDROCHLORIC ACID

Electromotive force measurements of cells without liquid junction, of the type Ag, AgCl, HCl + protein, H₂, have been made at 30°C. with the proteins gelatin, edestin, and casein in 0.1 M hydrochloric acid. The data are consistent with the assumptions of a constant combining capacity of each protein for hydrogen ion, no combination with chloride ion, and Failey''s principle of a linear variation of the logarithm of the mean activity coefficient of the acid with increasing protein concentration. The combining capacities for hydrogen ion so obtained are 13.4 x 10^–4 for edestin, 9.6 x 10^–4 for gelatin, and 8.0 x 10^–4 for casein, in equivalents of combined H⁺ per gm. of protein.     

AMPHOTERIC BEHAVIOR OF COMPLEX SYSTEMS : IV. NOTE ON THE ISOELECTRIC POINT AND IONIZATION CONSTANTS OF SULFANILIC ACID.

From the solubility minimum the value of the basic ionization constant of sulfanilic acid is shown to lie probably between the values 1.7 x 10^–15 and 3.2 x 10^–15. From solubility measurements the value of this same constant is shown to lie probably between 2.0 and 2.2 x 10^–15, and the isoelectric point of sulfanilic acid is thus at a cH of 0.056 or a pH of 1.25. From conductivity ratios the acid ionization constant of sulfanilic acid is shown to be 7.05 x 10^–4 at room temperature (21°C.). Calculations are made, from data published in preceding papers, of the ionization constants of glycine, K_a being 2.3 x 10^–10, and K_b being 2.2 x 10^–12.     

THE SOLUBILITY OF TYROSINE IN ACID AND IN ALKALI

Measurements have been made of the solubility at 25°C. of tyrosine in hydrochloric acid and in sodium hydroxide solutions varying from 0.001 to 0.05 M, and also in distilled water. The pH of the saturated solutions was measured with the hydrogen electrode. The following values for the ionization constants of tyrosine have been obtained from the measurements: k_b = 1.57 x 10^–12, k_a1 = 7.8 x 10^–10, k_a2 = 8.5 x 10^–11. The changes in solubility with pH can be satisfactorily explained by the use of these ionization constants.     

Muscle Contraction during Hyperpolarizing Currents in the Crab

Isolated muscle fibers from the motor legs of the crab Trichodactilus dilocarcinus were submitted to strong hyperpolarizing currents of varied intensities which produced tension during the current pulse. Threshold for tension was obtained with intensities of about 0.2 x 10^–5 A, changing E_m to ca. –150 mV (starting from a resting potential ofca. –80 mV). At the closure of the anodic square pulse, a second phase of tension usually appeared superimposed upon the one obtained during hyperpolarization. The first phase of tension increased with the increase of Ca⁺⁺ concentration in the bath. Sr⁺⁺ produced the same type of mechanical output as Ca⁺⁺. When added to the normal Ca⁺⁺ concentration, Ba⁺⁺ and Mn⁺⁺ in low concentrations (up to 21.5 mM) also increased the tension of this phase, but at higher concentrations they blocked both phases while Mg⁺⁺ did not alter the tension. Of all the divalent cations employed, only Sr⁺⁺ is capable of developing tension as a substitute for Ca⁺⁺ in the external media. Procaine administered in a dosage (5 x 10^–3 W/V)which would suppress the contracture due to caffeine (10 mM), did not modify the tension developed during the hyperpolarization. The preceding data indicate that the Ca⁺⁺ required for tension during hyperpolarization comes from sites which would differ from those usually postulated for tension due to depolarization in the muscle fibers of other crustaceans (American crayfish). Furthermore, the external source of Ca⁺⁺ appears to be one mainly implicated in the induction of tension due to inward current pulses.     

Effect of muscle mass on lactate formation during exercise in humans

To elucidate the mechanisms of lactate formation during submaximal exercise, eight men were studied during one- (1-LE) and two-leg (2-LE) exercise (approximately 11-min cycling) using the catheterization technique and muscle biopsies (quadriceps femoris muscle). The absolute exercise intensity and thus the energy demand for the exercising limb was the same [mean 114 (SEM 7) W] during both 1-LE and 2-LE. At the end of exercise partial pressure of O₂ and O₂ saturation in femoral venous blood were lower and arterial adrenaline and noradrenaline were higher during 2-LE than during 1-LE. Mean arterial blood lactate concentration increased to 10.8 (SEM 0.8) (2-LE) and 5.2 (SEM 0.4) mmol · 1^–1 (1-LE) after 10 min of exercise. The intramuscular metabolic response to exercise was attenuated during 1-LE [mean, lactate = 49 (SEM 9); glucose 6-P = 3.3 (SEM 0.3); nicotinamide adenine dinucleotide, reduced = 0.17 (SEM 0.02); adenosine 5-diphosphate 2.7 (SEM 0.1) mmol · kg dry mass^–1] compared to 2-LE [76 (SEM 6); 6.1 (SEM 0.7); 0.21 (SEM 0.02); 3.0 (SEM 0.1) mmol · kg dry mass^–1, respectively]. To elucidate whether the lower plasma adrenaline concentration could contribute to the attenuated metabolic response, additional experiments were performed on four of the eight subjects with infusion of adrenaline during 1-LE (1-LEE). Average plasma adrenaline concentration was increased during 1-LEE and reached 2–4 times higher levels than during 2-LE. Post-exercise muscle lactate and glucose 6-P contents were higher during 1-LEE than during 1-LE and were similar to those during 2-LE. Also, leg lactate release was elevated during 1-LEE versus 1-LE. It was concluded that during submaximal dynamic exercise the intramuscular metabolic response not only depended on the muscle power output, but also on the total muscle mass engaged. Plasma adrenaline concentrations and muscle oxygenation were found to be dependent upon the working muscle mass and both may have affected the metabolic response during exercise.     

FLUIDITY OF THE SURFACE OF CULTURED MUSCLE FIBERS : Rapid Lateral Diffusion of Marked Surface Antigens

Fluorescent antibody fragments of anti-muscle plasma membrane antibody bound as small fluorescent spots when applied by micropipetting to cultured myotubes. The spots were observed to enlarge with time. The rate of enlargement of fluorescent spots was greater when fragments were applied than when divalent antibody was used. It was also greater at 23°–25°C than at 0°–4°C. With glutaraldehyde-fixed cells no increase in the size of the spots was seen. The observations are consistent with the spread of fluorescent spots due to diffusion of surface protein antigens within the plane of a fluid membrane. From measurements of spot size against time, a diffusion constant of 1–3 x 10^-9 cm² s^-1 can be calculated for muscle plasma membrane proteins of mol wt approximately 200,000. This value is consistent with other observations on the diffusion of surface antigens and of labeled lipid molecules in synthetic and natural membranes.     

THE BEHAVIOR OF THE NUCLEIC ACIDS DURING THE EARLY DEVELOPMENT OF THE SEA URCHIN EGG (ARBACIA)

The authors wish to correct an error in the paper "The behavior of the nucleic acids during the early development of the sea urchin egg (Arbacia)" (J. Gen. Physiol., 1947–48, 31, 203). Owing to an oversight, the figures for the amounts of various P fractions in a single Arbacia egg have been erroneously expressed in γ x 10^–3 units (Tables I and II, page 205; the last two lines of page 206). The figures should have been expressed in γ x 10^–5 units. Thus, the fertilized Arbacia egg contains an average of 20 γ x 10^–5 ribonucleic acid P and 0.7 to 1 γ x 10^–5 desoxyribonucleic acid P.     

SOLUBILITY OF MUSTARD GAS [BIS (β-CHLOROETHYL) SULFIDE] IN WATER,MOLAR SODIUM CHLORIDE,AND IN SOLUTIONS OF DETERGENTS

1. The solubility of mustard (H) in water and in molar sodium chloride was found to be 5.8 x 10^–3 molar and 3.2 x 10^–3 molar respectively or 0.92 mg. per ml. and 0.5 mg. per ml. Solubility curves have been drawn and the usefulness of this method in examining the homogeneity of H preparations as well as in establishing their solubility, is discussed. 2. Certain detergents increase the solubility of H in water. The solubility was found to increase with the concentration of detergent. 3. Many detergents were found to affect the interfacial tension between H and water so that with slight agitation liquid H breaks up into minute droplets. This in turn greatly accelerates the rate of solution.     

Plasma ATP concentration and venous oxygen content in the forearm during dynamic handgrip exercise

Background  

It has been proposed that adenosine triphosphate (ATP) released from red blood cells (RBCs) may contribute to the tight coupling between blood flow and oxygen demand in contracting skeletal muscle. To determine whether ATP may contribute to the vasodilatory response to exercise in the forearm, we measured arterialised and venous plasma ATP concentration and venous oxygen content in 10 healthy young males at rest, and at 30 and 180 seconds during dynamic handgrip exercise at 45% of maximum voluntary contraction (MVC).     

THE ACTIVATION OF STARFISH EGGS BY ACIDS : II. THE ACTION OF SUBSTITUTED BENZOIC ACIDS AND OF BENZOIC AND SALICYLIC ACIDS AS INFLUENCED BY THEIR SALTS.

1. Comparison of the rates of activation of unfertilized starfish eggs in pure solutions of a variety of parthenogenetically effective organic acids (fatty acids, carbonic acid, benzoic and salicylic acids, chloro- and nitrobenzoic acids) shows that solutions which activate the eggs at the same rate, although widely different in molecular concentration, tend to be closely similar in C_H. The dissociation constants of these acids range from 3.2 x 10^–7 to 1.32 x 10^–3. 2. In the case of each of the fourteen acids showing parthenogenetic action the rate of activation (within the favorable range of concentration) proved nearly proportional to the concentration of acid. The estimated C_H of solutions exhibiting an optimum action with exposures of 10 minutes (at 20°) lay typically between 1.1 x 10^–4 M and 2.1 x 10^–4 M (pH = 3.7–3.96), and in most cases between 1.6 x 10^–4 M and 2.1 x 10^–4 M (pH = 3.7–3.8). Formic acid (C_H = 4.2 x 10^–4 M) and o-chlorobenzoic acid (C_H = 3.5 x 10^–4 M) are exceptions; o-nitrobenzoic acid is ineffective, apparently because of slow penetration. 3. Activation is not dependent on the penetration of H ions into the egg from without, as is shown by the effects following the addition of its Na salt to the solution of the activating acid (acetic, benzoic, salicylic). The rate of activation is increased by such addition, to a degree indicating that the parthenogenetically effective component of the external solution is the undissociated free acid. Apparently the undissociated molecules alone penetrate the egg freely. It is assumed that, having penetrated, they dissociate in the interior of the egg, furnishing there the H ions which effect activation. 4. Attention is drawn to certain parallels between the physiological conditions controlling activation in the starfish egg and in the vertebrate respiratory center.     

MITOTIC AND NONMITOTIC MULTIPLE-LAYERED PERFUSION CULTURES

Cell types in addition to those previously described (Kruse et al. 1963. J. Nat. Cancer Inst. 31:109; Kruse and Miedema. 1965. J. Cell Biol. 27:273) were found to form multiple-layered cultures by perfusion-culture technique. Dense populations containing 43 x 10⁶ embryonic rat muscle (NF-ER) cells, 23 x 10⁶ diploid human tonsillar (NF-JAM) cells, 77 x 10⁶ human pleural effusion isolate (RPMI 2650) cells, 35 x 10⁶ embryonic diploid human lung (Flow 2000) cells, 21 x 10⁶ bovine lung (FB4BM) cells, 108 x 10⁶ bat lung (Tb1Lu) cells, and 81 x 10⁶ SV-40 virus-transformed embryonic diploid human lung (WI-38VA13A) cells were obtained in 6–14 days from dilute inocula in T-60 or T-75 flasks; these were equivalent to about 4, 3, 3, 4, 2, 4, and eight monolayers, respectively. Perfusion of an NF-ER culture for 6 wk with medium plus 10% whole calf serum yielded a cell density equivalent to 12 monolayers (140 x 10⁶ cells per T-75 flask). This culture exhibited random labeling of nuclei from bottom to top after pulsing for 90 min with thymidine-³H. Medium plus 0.1% serum maintained NF-JAM cultures at constant viable cell numbers with virtual absence of thymidine-³H labeling. Similar results were obtained with WI-38 cultures, but WI-38VA13A cells continued active DNA synthesis and mitosis in medium with 0.1% serum to form 16–20 layers of cells (191–239 x 10⁶ cells per T-75 flask) in 27 days. WI-38VA13A cells ceased proliferation and became nonviable rapidly in serumless medium.     

PERMEABILITY OF ERYTHROCYTES TO RADIOACTIVE POTASSIUM

The diffusion coefficients for the exchange of potassium across the membrane of erythrocytes of humans, rats, and rabbits have been determined by the use of artificially radioactive potassium, both into and out of the erythrocytes both in vitro and in vivo. The diffusion coefficients found in minutes^–1 were 0.2 to 0.25 x 10^–3 for human, 0.32 to 0.665 x 10^–3 for rabbits, and 1.0 x 10^–3 for rat erythrocytes. Rabbit erythrocytes appear to be more permeable in vivo. Reasons are advanced to explain the failure of earlier workers to demonstrate appreciable exchange of potassium in erythrocytes.     

Intracellular Calcium Binding and Release in Frog Heart

The capacities and affinities of intracellular calcium-binding sites have been studied in frog ventricles, in which the concentration of Ca⁺⁺ in the sarcoplasm can be controlled as a result of treatment with EDTA. The total calcium content of calcium-depleted and nondepleted muscles at rest and muscles generating considerable tension was 0.8, 1.4, and 5.4 µmol/g of muscle, respectively. Net movement of calcium into or out of the cells occurred without change in tension when the sarcoplasmic concentration of Ca⁺⁺ was either of two values, less than 10^-7 M or approximately 5 x 10^-7 M. These data can be explained by the presence of two groups of intracellular calcium sinks which compete with the contractile proteins, one with a capacity of about 0.6 µmol/g and an affinity constant greater than 10⁷ M^-1 and a second with a capacity of 4.0 µmol/g and an affinity constant of about 2 x 10⁶ M^-1. The higher affinity calcium is released by anoxia, oligomycin, or abrupt changes in sarcoplasmic Ca⁺⁺. Muscles soaked in Sr-Ringer's contain electron densities in the sarcoplasmic reticulum and to a lesser extent in the mitochondria.     

End Criteria for Reaching Maximal Oxygen Uptake Must Be Strict and Adjusted to Sex and Age: A Cross-Sectional Study

Objective

To describe different end criteria for reaching maximal oxygen uptake (VO_2max) during a continuous graded exercise test on the treadmill, and to explore the manner by which different end criteria have an impact on the magnitude of the VO_2max result.

Methods

A sample of 861 individuals (390 women) aged 20–85 years performed an exercise test on a treadmill until exhaustion. Gas exchange, heart rate, blood lactate concentration and Borg Scale_6–20 rating were measured, and the impact of different end criteria on VO_2max was studied;VO₂ leveling off, maximal heart rate (HR_max), different levels of respiratory exchange ratio (RER), and postexercise blood lactate concentration.

Results

Eight hundred and four healthy participants (93%) fulfilled the exercise test until voluntary exhaustion. There were no sex-related differences in HR_max, RER, or Borg Scale rating, whereas blood lactate concentration was 18% lower in women (P<0.001). Forty-two percent of the participants achieved a plateau in VO₂; these individuals had 5% higher ventilation (P = 0.033), 4% higher RER (P<0.001), and 5% higher blood lactate concentration (P = 0.047) compared with participants who did not reach a VO₂ plateau. When using RER ≥1.15 or blood lactate concentration ≥8.0 mmol•L^–1, VO_2max was 4% (P = 0.012) and 10% greater (P<0.001), respectively. A blood lactate concentration ≥8.0 mmol•L^–1 excluded 63% of the participants in the 50–85-year-old cohort.

Conclusions

A range of typical end criteria are presented in a random sample of subjects aged 20–85 years. The choice of end criteria will have an impact on the number of the participants as well as the VO_2max outcome. Suggestions for new recommendations are given.     

STIMULATION BY MINERAL AND FATTY ACIDS IN THE BARNACLE BALANUS BALANOIDES

1. Stimulation in the rock barnacle Balanus balanoides by hydrochloric, sulfuric, and nitric acids, and by the first seven members of the normal aliphatic acid series has been studied. The hydrogen ion concentrations of the solutions tested varied from 3.2 x 10^–8 to 5.889 x 10^–6. The criterion of response was percentage closure in groups of individuals, recorded at 1 minute intervals until maximum closure occurred. 2. The intensity of stimulation by these acids is proportional to the effects of two forces, one related to the change in the (H⁺), and the other to the field of force around the anion of the acid added to the environment. 3. A preliminary interpretation of the results led to the development of the following expression which fits approximately the data obtained at the end of 4 minutes: Per cent closure = 100 – 100e ^{–0.1z+(0.003125)2–0.1z+(0.003125)2n(z–0.4)} where z is the (H⁺) x 10⁷ and n is the number of carbon atoms (if present) in the anion of the acid. This equation assumes that the anions of the mineral acids enter into the reaction stoichiometrically, and emphasizes the difference in the fields of force around the anion of the fatty acids, a difference which is correlated with the length of the carbon chain. 4. A further analysis of the data revealed the presence of three or more receptor groups which appeared to be differentially affected by forces originating from the anions of the acids. 5. The order of stimulating efficiency for the mineral acids was found to be: HCl>H₂SO₄>HNO₃. 6. The order of stimulating efficiency for the fatty acids was found to be: heptylic>caproic>valeric>butyric = acetic>propionic = formic.     

The Impact of Fish and the Commercial Marine Harvest on the Ocean Iron Cycle

Although iron is the fourth most abundant element in the Earth''s crust, bioavailable iron limits marine primary production in about one third of the ocean. This lack of iron availability has implications in climate change because the removal of carbon dioxide from the atmosphere by phytoplankton requires iron. Using literature values for global fish biomass estimates, and elemental composition data we estimate that fish biota store between 0.7–7×10¹¹ g of iron. Additionally, the global fish population recycles through excretion between 0.4–1.5×10¹² g of iron per year, which is of a similar magnitude as major recognized sources of iron (e.g. dust, sediments, ice sheet melting). In terms of biological impact this iron could be superior to dust inputs due to the distributed deposition and to the greater solubility of fecal pellets compared to inorganic minerals. To estimate a loss term due to anthropogenic activity the total commercial catch for 1950 to 2010 was obtained from the Food and Agriculture Organization of the United Nations. Marine catch data were separated by taxa. High and low end values for elemental composition were obtained for each taxonomic category from the literature and used to calculate iron per mass of total harvest over time. The marine commercial catch is estimated to have removed 1–6×10⁹ g of iron in 1950, the lowest values on record. There is an annual increase to 0.7–3×10¹⁰ g in 1996, which declines to 0.6–2×10¹⁰ g in 2010. While small compared to the total iron terms in the cycle, these could have compounding effects on distribution and concentration patterns globally over time. These storage, recycling, and export terms of biotic iron are not currently included in ocean iron mass balance calculations. These data suggest that fish and anthropogenic activity should be included in global oceanic iron cycles.     

Viral Load Levels Measured at Set-Point Have Risen Over the Last Decade of the HIV Epidemic in the Netherlands

Background

HIV-1 RNA plasma concentration at viral set-point is associated not only with disease outcome but also with the transmission dynamics of HIV-1. We investigated whether plasma HIV-1 RNA concentration and CD4 cell count at viral set-point have changed over time in the HIV epidemic in the Netherlands.

Methodology/Principal Findings

We selected 906 therapy-naïve patients with at least one plasma HIV-1 RNA concentration measured 9 to 27 months after estimated seroconversion. Changes in HIV-1 RNA and CD4 cell count at viral set-point over time were analysed using linear regression models. The ATHENA national observational cohort contributed all patients who seroconverted in or after 1996; the Amsterdam Cohort Studies (ACS) contributed seroconverters before 1996. The mean of the first HIV-1 RNA concentration measured 9–27 months after seroconversion was 4.30 log₁₀ copies/ml (95% CI 4.17–4.42) for seroconverters from 1984 through 1995 (n = 163); 4.27 (4.16–4.37) for seroconverters 1996–2002 (n = 232), and 4.59 (4.52–4.66) for seroconverters 2003–2007 (n = 511). Compared to patients seroconverting between 2003–2007, the adjusted mean HIV-1 RNA concentration at set-point was 0.28 log₁₀ copies/ml (95% CI 0.16–0.40; p<0.0001) and 0.26 (0.11–0.41; p = 0.0006) lower for those seroconverting between 1996–2002 and 1984–1995, respectively. Results were robust regardless of type of HIV-1 RNA assay, HIV-1 subtype, and interval between measurement and seroconversion. CD4 cell count at viral set-point declined over calendar time at approximately 5 cells/mm³/year.

Conclusion

The HIV-1 RNA plasma concentration at viral set-point has increased over the last decade of the HIV epidemic in the Netherlands. This is accompanied by a decreasing CD4 cell count over the period 1984–2007 and may have implications for both the course of the HIV infection and the epidemic.