细针穿刺甲状腺肿瘤组织细胞DNA,RNA含量分析

本组资料应用流式细胞分析技术对１１２例甲状腺肿瘤细针穿刺组织细胞的ＤＮＡ、ＲＮＡ含量进行了定量研究，结果细针穿刺吸取组织行流式细胞ＤＮＡ、ＲＮＡ分析的成功率为９８．２％。１１０例术前针吸新鲜组织与术后石蜡包埋组织ＤＮＡ、ＲＮ含量无统计学差别（Ｐ＞０．０５）。ＤＮＡ异倍体可见于良性甲状腺肿瘤，随访提示，良性甲状腺肿瘤ＤＮＡ异倍体与术后预后无关。     

小儿恶性肿瘤细胞的DNA含量及其临床意义

目的：探讨小儿恶性肿瘤ＤＮＡ含量及其与临床预后的关系。方法：用流式细胞仪对６２例小儿恶性肿瘤细胞的ＤＮＡ含量进行测定，并分析其中３５例的细胞增殖状况及其与临床分期的关系。结果：ＤＮＡ组方图呈二倍体的肿瘤１４例，异倍体的肿瘤４８例（检出率７７．４％）；Ⅲ～Ⅳ期肿瘤的异倍体检出率高达９７．４％；ＤＮＡ指数（ＤＩ）和倍性与肿瘤的分型无关。结论：测定结果应与临床、病理资料结合，对预后作出准确判断     

B超结合血清CA－125测定诊断卵巢恶性肿瘤

１２１例卵巢肿瘤患者手术后经病理学诊断，本文对他们的术前Ｂ超结果和ＣＡ－１２５浓度进行分析评价。Ｂ超对卵巢恶性肿瘤的检测灵敏度为８５％，特异性８９％。ＣＡ－１２５阳性界值为４５Ｕ／ｍｌ时，诊断恶性卵巢肿瘤的灵敏度为９７％，特异性８２％，两者联合使用的特异性９８％，联合阳性预测值（ＰＰＶ）为９５％；当ＣＡ－１２５的阳性界值定为６５Ｕ／ｍｌ时，特异性为９９％，ＰＰＶ９８％。因此，Ｂ超和ＣＡ－１２５联合检测，为卵巢恶性肿瘤的诊断提供了一个有力的指标。     

检测丁型肝炎病毒RNA打点杂交法的建立及其应用

经缺口平移法以ａ－３２Ｐ－ｄＣＴＰ标记１．０ｋｂ的丁型肝炎病毒（ＨＤＶ）ｃＤＮＡ片段为探针，采用蛋白酶Ｋ直接从血清中提取ＨＤＶＲＮＡ，建立了检测血清中ＨＤＶＲＮＡ的打点杂交法，其灵敏性可达１ｐｇ水平，与乙型肝炎病毒（ＨＢＶ）ＤＮＡ无交叉杂交反应；并应用于检测我国５９４９份ＨＢｓＡｇ阳性血清中的ＨＤＶＲＮＡ，共检出１７６份ＨＤＶＲＮＡ阳性，检出率为２．９５％。     

地高辛标记大鼠下丘脑生长激素释放因子cRNA探针的制备及应用

制备地高辛标记大鼠下丘脑释放因子ｃＲＮＡ探针；方法；大鼠下丘脑生长激素释放因子的重组质粒ｃＤＮＡ＝ＰＧＥＭ４经转化扩增后，用碱性裂妥法获取质粒ｃＤＮＲ并纯化。用限制性内切酶ＥＣＯＲＩ酶切，以线性ｃＤＮＡ为模板，在Ｔ７和Ｓｐ６ＲＮＡ聚合酶作用下，采用体外转录法分别合成地高辛素标记的大鼠下丘脑生激素释放因子ｃＲＮＡ和ＲＮＡ探针。     

中国湖南株丁型肝炎病毒cDNA（443－870）片段的克隆和序列分析

取１例湖南丁型肝炎病毒（ＨＤＶ）ＲＮＡ阳性血清，经强变性剂法抽提ＨＤＶＲＮＡ，逆转录－聚合酶链反应（ＲＴ－ＰＣＲ）扩增，扩增的ＨＤＶＣＤＮＡ片段重组到ｐＵＣ１８质粒中，获得了中国湖南株ＨＤＶＣＤＮＡ（４３３－８７０）片段克隆，ＤＮＡ测序结果显示，该片段（４３８ｈｐ长，包括一端ＰＣＲ引物２５ｈｐ）与已知的美国、意大利、法国、诺鲁和中国台湾５株ＨＤＶｃＤＮＡ相同片段比较，同源性分别为９１．３％，９４．５％，９１．３％，８４．５％和８９．３％，其中与ＨＤＶＲＮＡ复制密切相关的第一个高度保守区与美国株、意大利株和法国株完全同源。     

原位杂交法检测肝组织中丁型和乙型肝炎病毒核酸

利用国外引进的重组质粒获得纯化基因片段，分别以随机引物法和ＰＣＲ法制备地高辛素标记的ＨＢＶＤＮＡ探针和ＨＤＶｃＤＮＡ探针。用原位杂交法检测了石蜡包埋的肝组织切片ＢＶＤＮＡ和ＨＤＶＲＮＡ。４９例感染肝组织分为两组：丁肝组２３例；单纯乙肝组２６例，ＨＢＶＤＮＡ的检出率丁肝组（７８．２６％）与乙肝组（７６．９２％）无统计学差异；而ＨＤＶＮＡ的检出率丁肝组（６０．８７％）明显高于乙肝组（１５．３８％）。ＨＢＶＤＮＡ可见于受染肝细胞的胞核或胞浆内，而ＨＤＶＲＮＡ绝大部分见于肝细胞胞核。两种病毒核酸阳性细胞在肝组织中的分布特点大致相同：弥漫或散在地分布于肝小叶或假小叶内，或局灶性分布于小叶周边。ＨＤＶＲＮＡ阳性的肝组织都或多或少地同时存在ＨＢＶＤＮＡ。同一例肝组织中，ＨＢＶＤＮＡ阳性细胞从数量和颗粒密度上似略高于ＨＤＶＲＮＡ。将乙肝组和丁肝组两组病人肝内ＨＢ－ｓＡｇ、ＨＢｃＡｇ和ＨＢＶＤＮＡ及血清ＨＢｅＡｇ作了比较，各指标阳性率虽有差异，但均无统计学意义。因此，未发现ＨＤＶ感染对ＨＢＶ的复制有明显抑制作用。此结果对以往用血清学或免疫组化方法对ＨＤＶ的研究有所补充和深入，亦可为研究其它类型病毒性肝炎之间的重叠感染所借鉴。     

涎腺粘液表皮样癌DNA指数与分化及淋巴结转移的关系

目的：研究涎腺粘液表皮样癌细胞ＤＮＡ指数及ＤＮＡ倍体情况，评估其与粘液表皮样癌癌细胞分化及淋巴结转移的关系。方法：收集４０例涎腺粘淮有皮样癌，用ＩＣＭ对其进行ＤＮＡ含量测定。结果：正常涎腺导管上皮及良性肿瘤细胞核ＤＩ均值分别为１．０２７和１．０２２，均为正常二倍体；而４０例涎腺粘液表皮样癌癌细胞核Ｄ一值为１．３８３，异倍体出现率为６２．５％；随着粘液表皮样癌癌细胞分化程度降低，ＤＩ值及ＤＮＡ异倍体     

用聚合酶链反应检测人肝细胞癌乙型肝炎病毒DNA和丙型肝炎病毒RNA

为研究乙型肝炎病毒ＤＮＡ（ＨＢＶＤＮＡ）和丙型肝炎病毒ＲＮＡ（ＨＣＶＤＮＡ）与肝细胞癌的关系，用聚合酶链反应（ＰＣＲ）和巢式ＰＣＲ（ｎｅｓｔｅｄ－ＰＣＲ）分别检测４２例肝肿瘤组织中ＨＢＶＤＮＡ和ＨＣＶＲＮＡ。结果：１例胆管细胞癌组织ＨＢＶＤＮＡ和ＨＣＶＲＮＡ均阳性，１例胆管囊腺瘤ＨＢＶＤＮＡ阳性。４０例肝细胞癌组织中，单纯ＨＢＶＤＮＡ阳性１９例，单纯ＨＣＶＲＮＡ阳性３例，二者均阳性１０例。ＨＢＶＤＮＡ阳性率７２．５％，ＨＣＶＲＮＡ阳性率３２．５％。ＨＢＶＤＮＡ和ＨＣＶＲＮＡ感染与肝癌组织学分型无关；且肝细胞癌中ＨＣＶ感染与ＨＢＶ未见相关。结果提示，我国ＨＢＶ感染仍是引起肝细胞癌的主要原因。但由于肝细胞癌患者中ＨＣＶ的感染率也较高，且有上升趋势，因此ＨＣＶ可能也是肝细胞癌发生的重要原因之一。     

斑点免疫金银染色检测抗双链DNA抗体

从牛胸腺ＤＮＡ制备双链ＤＮＡ溶液，点于混合纤维素酯微孔滤膜，进行斑点免疫金银染色（Ｄｏｔ－ＩＧＳＳ），以检测抗双链ＤＮＡ抗体（ＡｄｓＤＮＡ）。在３１６份所测血清中此法检出的ＡｄｓＤＮＡ对ＳＬＥ的敏感性、特异性分别是６９．４％、９６．８％，与平行对照的ＥＳＮ／ＡＥＳ－ＩＧＳＳ、ＲＩＡ、ＥＬＩＳＡ等法相比差异无显著性（Ｐ＞０．０５），符合率均＞９２％。此法操作方便、结果观察简单明了，更为实用。     

Opportunities and challenges in the prevention and control of cancer and other chronic diseases: children's diet and nutrition and weight and physical activity

OBJECTIVE: The purpose of this article is to review the role of behavioral research in disease prevention and control, with a particular emphasis on lifestyle- and behavior-related cancer and chronic disease risk factors--specifically, relationships among diet and nutrition and weight and physical activity with adult cancer, and tracking developmental origins of these health-promoting and health-compromising behaviors from childhood into adulthood. METHOD: After reviewing the background of the field of cancer prevention and control and establishing plausibility for the role of child health behavior in adult cancer risk, studies selected from the pediatric published literature are reviewed. Articles were retrieved, selected, and summarized to illustrate that results from separate but related fields of study are combinable to yield insights into the prevention and control of cancer and other chronic diseases in adulthood through the conduct of nonintervention and intervention research with children in clinical, public health, and other contexts. RESULTS: As illustrated by the evidence presented in this review, there are numerous reasons (biological, psychological, and social), opportunities (school and community, health care, and family settings), and approaches (nonintervention and intervention) to understand and impact behavior change in children's diet and nutrition and weight and physical activity. CONCLUSIONS: Further development and evaluation of behavioral science intervention protocols conducted with children are necessary to understand the efficacy of these approaches and their public health impact on proximal and distal cancer, cancer-related, and chronic disease outcomes before diffusion. It is clear that more attention should be paid to early life and early developmental phases in cancer prevention.     

Pain and Effusion and Quadriceps Activation and Strength

Context:

Quadriceps dysfunction is a common consequence of knee joint injury and disease, yet its causes remain elusive.

Objective:

To determine the effects of pain on quadriceps strength and activation and to learn if simultaneous pain and knee joint effusion affect the magnitude of quadriceps dysfunction.

Design:

Crossover study.

Setting:

University research laboratory.

Patients or Other Participants:

Fourteen (8 men, 6 women; age = 23.6 ± 4.8 years, height = 170.3 ± 9.16 cm, mass = 72.9 ± 11.84 kg) healthy volunteers.

Intervention(s):

All participants were tested under 4 randomized conditions: normal knee, effused knee, painful knee, and effused and painful knee.

Main Outcome Measure(s):

Quadriceps strength (Nm/kg) and activation (central activation ratio) were assessed after each condition was induced.

Results:

Quadriceps strength and activation were highest under the normal knee condition and differed from the 3 experimental knee conditions (P < .05). No differences were noted among the 3 experimental knee conditions for either variable (P > .05).

Conclusions:

Both pain and effusion led to quadriceps dysfunction, but the interaction of the 2 stimuli did not increase the magnitude of the strength or activation deficits. Therefore, pain and effusion can be considered equally potent in eliciting quadriceps inhibition. Given that pain and effusion accompany numerous knee conditions, the prevalence of quadriceps dysfunction is likely high.Key Words: arthrogenic muscle inhibition, central activation failure, voluntary activation, muscles

Key Points

• Knee pain and effusion resulted in arthrogenic muscle inhibition and weakness of the quadriceps.
• The simultaneous presence of pain and effusion did not increase the magnitude of quadriceps dysfunction.
• To reduce arthrogenic muscle inhibition and improve muscle strength, clinicians should employ interventions that target removing both pain and effusion.

Quadriceps weakness is a common consequence of traumatic knee joint injury^1,2 and chronic degenerative knee joint conditions.^3,4 Arthrogenic muscle inhibition (AMI), a neurologic decline in muscle activation, results in quadriceps weakness and hinders rehabilitation by preventing gains in strength.⁵ The inability to reverse AMI and restore muscle function can lead to decreased physical abilities,⁶ biomechanical deficits,⁷ and possibly reinjury.⁵ Furthermore, researchers^8,9 have suggested that quadriceps weakness resulting from AMI may place patients at risk for developing osteoarthritis in the knee. In light of the substantial influence of quadriceps AMI on these clinically relevant outcomes, we need to improve our understanding of the factors that contribute to this neurologic decline in muscle activity so efforts to target and reverse it can be implemented and gains in strength can be achieved more easily.Joint injury and disease are accompanied by numerous sequelae (ie, pain, swelling, tissue damage, inflammation), so ascertaining which one ultimately leads to neurologic muscle dysfunction is difficult. Whereas a joint effusion can result in AMI,^10–12 the effects of pain are less understood despite many clinicians attributing AMI to pain. Using techniques that introduce knee pain without accompanying injury may provide insights into the role of pain in eliciting AMI.The degree of knee joint damage may play a role in the quantity of AMI that manifests. Hurley et al^13,14 demonstrated that quadriceps AMI, measured using an interpolated-twitch technique, was greater in patients with extensive traumatic knee injury (eg, fractured tibial plateau, ruptured medial collateral ligament, and medial meniscectomy) than patients with isolated joint trauma (ie, isolated anterior cruciate ligament [ACL] rupture). Similarly, patients with more knee joint symptoms (ie, greater number of symptoms and increased severity of symptoms) may present with greater magnitudes of quadriceps inhibition. Recently, investigators¹⁵ have suggested that patients with more pain display less quadriceps strength, supporting this tenet. Given that effusion and pain often present simultaneously with joint injuries and diseases, such as ACL injury and osteoarthritis, examining both the isolated and cumulative effects of these sequelae appears warranted to determine if they influence the magnitude of muscle inhibition.Experimental joint-effusion and pain models are safe and effective experimental methods that allow for the isolated examination of their effects on muscle function. The effusion model, whereby sterile saline is injected directly into the knee joint capsule,⁷ produces a clinically relevant magnitude of the joint effusion that may be present with traumatic injury. Effusion is thought to activate group II afferents responding to stretch or pressure,^16–18 which in turn may facilitate group Ib interneurons and result in quadriceps AMI.⁵ The pain model involves injecting hypertonic saline into the infrapatellar fat pad to produce anteromedial knee pain similar to that described in patients with patellofemoral pain syndrome.¹⁹ Pain is considered to initiate AMI through activation of group III and IV afferents that act as nocioceptors to signal damage or potential damage to joint structures.^16–18 The firing of these afferents then may lead to facilitation of group Ib interneurons, the flexion reflex, or the gamma loop, ultimately resulting in quadriceps inhibition.²⁰ Thus, these models allow us to create symptoms that are associated with knee injury and have the added benefit of providing a way to examine their effects in isolation.Therefore, the purpose of our study was to determine the effects of pain on quadriceps strength and activation and to learn if simultaneous pain and knee joint effusion would affect the magnitude of quadriceps dysfunction. We hypothesized that pain alone would result in quadriceps inhibition and that the magnitude of inhibition would be greater when effusion and pain were present simultaneously.     

Postinfectious immunodeficiency and autoimmunity: pathogenic and clinical values and implications

Autoimmunity is still a mystery of clinical immunology and medicine as a whole. The etiology and pathogenesis of autoimmune disorders remain unclear and, thus, are assessed as a balance between hereditary predisposition, triggering factors and the appearance of autoantibodies and/or self-reactive T cells. Among the immunological armamentarium, molecular mimicry, based on self-reactive T- and B-cell activation by cross-reactive epitopes of infectious agents, is of special value. Hypotheses regarding the possible involvement of molecular mimicry in the development of postinfectious autoimmunity are currently very intriguing. They provide new approaches for identifying etiological agents that are associated with postinfectious autoimmunity, paired microbial- and tissue-linked epitopes targeted for autoimmune reaction determination, postinfectious autoimmunity pathogenesis recognition and specific prevention, and therapy for autoimmune disorder development.     

Beta-endorphin and drug-induced reward and reinforcement

Although drugs of abuse have different acute mechanisms of action, their brain pathways of reward exhibit common functional effects upon both acute and chronic administration. Long known for its analgesic effect, the opioid beta-endorphin is now shown to induce euphoria, and to have rewarding and reinforcing properties. In this review, we will summarize the present neurobiological and behavioral evidences that support involvement of beta-endorphin in drug-induced reward and reinforcement. Currently, evidence supports a prominent role for beta-endorphin in the reward pathways of cocaine and alcohol. The existing information indicating the importance of beta-endorphin neurotransmission in mediating the reward pathways of nicotine and THC, is thus far circumstantial. The studies described herein employed diverse techniques, such as biochemical measurements of beta-endorphin in various brain sites and plasma, and behavioral measurements, conducted following elimination (via administration of anti-beta-endorphin antibodies or using mutant mice) or augmentation (by intracerebral administration) of beta-endorphin. We suggest that the reward pathways for different addictive drugs converge to a common pathway in which beta-endorphin is a modulating element. beta-Endorphin is involved also with distress. However, reviewing the data collected so far implies a discrete role, beyond that of a stress response, for beta-endorphin in mediating the substance of abuse reward pathway. This may occur via interacting with the mesolimbic dopaminergic system and also by its interesting effects on learning and memory. The functional meaning of beta-endorphin in the process of drug-seeking behavior is discussed.     

PTEN与信号转导及肿瘤

ＴＥＮ[1] (ｐｈｏｓｐｈａｔａｓｅａｎｄｔｅｎｓｉｎｈｏｍｏｌｏｇｙｄｅｌｅｔｅｄｏｎｃｈｒｏｍｏｓｏｍｅｔｅｎ)又名ＭＭＡＣ1 [2 ] (ｍｕｔａｔｅｄｉｎｍｕｔｉｐｌｙａｄａｎｃｅｄｃａｎｃｅｒ 1 )和ＴＥＰ1 [3 ] (ＴＧＦ -βｒｅｇｕｌａｔｅｄａｎｄｅｐｉｔｈｅｌｉａｌｃｅｌｌ -ｒｉｃｈｅｄｐｈｏｓｐｈａｔａｓｅ 1 ) (以下均称为ＰＴＥＮ) ,是 1 997年由 3个研究小组先后发现的一个具有双特异磷酸酶活性的抑癌基因。ＰＴＥＮ基因异常广泛存在于人类多种恶性肿瘤 ,如恶性神经胶质瘤、前列腺癌、子宫内膜癌、黑色素瘤等…     

Tobacco and alcohol and the risk of head and neck cancer

Summary We carried out two case-control studies on the relative risk of head and neck cancer in association with tobacco and alcohol consumption. The first study carried out at the ENT Department of the University hospitals of Heidelberg and Giessen (FRG) comprised 200 male patients with squamous cell cancer of the head and neck and 800 control subjects matched for sex, age, and residential area (1:4 matching design). Of the tumour patients, 4.5% had never smoked, in contrast to 29.5% of the control group. The average tobacco and alcohol consumption of the patients was approximately twice as high as in the control subjects. The highest alcohol and tobacco consumption was observed in patients suffering from oropharyngeal cancer. Tobacco and alcohol increased the risk of head and neck cancer in a dose-dependent fashion and acted as independent risk factors. In heavy smokers (> 60 pack-years) a relative risk of 23.4 (alcohol adjusted) was calculated. Combined alcohol and tobacco consumption showed a synergistic effect. The risk ratio increased more in a multiplicative than in an additive manner. Oral and laryngeal cancer were associated with the highest tobacco-associated risk values. The highest ethanol-associated risk values were associated with oropharyngeal and laryngeal cancer. The second study was carried out at the ENT Department of the University of Heidelberg on 164 males with squamous cell carcinoma of the larynx and 656 control subjects matched for sex, age and residential area (1:4 matching design). Of the cases, 4.2% had never smoked, compared with 28.5% of the control subjects. The risk of laryngeal cancer by tobacco consumption was dose dependent, reaching a maximum value of 9.1 (adjusted for alcohol) for a consumption of more than 50 tobacco-years (TY). The relative risk of laryngeal cancer associated with alcohol intake was also dose dependent, reaching a value of 9.0 (adjusted for tobacco) for a mean daily consumption of more than 75 g alcohol. An analysis of subsite specific risks showed that heavy smokers (> 50 TY) carried a nearly ten times higher risk of supraglottic cancer than of glottic cancer. The risk of supraglottic cancer from alcohol consumption was also higher than that of glottic cancer.     

Muscle strength and serum testosterone,cortisol and SHBG concentrations in middle-aged and elderly men and women

Forty healthy males (M) and females (F) divided into two different age groups i.e. M50 years (range 44–57; n= 9), F50 years (range 43–54; n= 9), M70 years (range 64–73; n= 11) and F70 years (range 63–73; n= 11) volunteered as subjects for examination of muscle cross-sectional area (CSA) and maximal voluntary isometric force production characteristics of the leg extensor muscles and serum androgen and sex hormone binding globulin (SHBG) concentrations. The CSA in the male groups was greatly larger (P < 0.01) than in the female groups and both elderly groups demonstrated slightly (n.s.) smaller values in the CSA than the two middle-aged groups. Maximal force of 2854 ± 452 N in M50 was greater (P < 0.05) than that of 2627 ± 752 N recorded for F50 as well as the force of 2787 ± 843 in M70 was greater (P < 0.001) than that of 1849 ± 295 recorded for F70. The force between F50 and F70 differed significantly (P < 0.05) from each other. The maximal rate of force production in M50 was greater (P < 0.01) than in F50 as well as in M70 greater (P < 0.001) than in F70. Both middle-aged groups demonstrated greater (P < 0.05) values than the respective elderly groups of the same sex. The individual values in the CSA correlated with the values in maximal force both in the middle-aged subjects (r= 0.66; P < 0.01) and in the elderly subjects (r= 0.69; P < 0.01). The mean concentration of serum testosterone in M50 was slightly (n.s.) greater than in M70 and in F50 significantly (P < 0.05) greater than in F70. Serum SHBG levels were lower in the males (P < 0.01) than in the females and serum testosterone/SHBG ratio in M70 and in F70 were lower (P < 0.05) than in M50 and in F50, respectively. In the females significant positive correlations were observed between the individual values in serum testosterone concentration and the values both in the CSA (r= 0.46; P < 0.05) and in maximal force (r= 0.62; P < 0.01) as well as between serum testosterone/SHBG ratio and both the CSA (r= 0.55; P < 0.05) and maximal force (r= 0.68; P < 0.01). The present results imply that the decreasing basal level of blood testosterone over the years in aging people, especially in females, may lead to decreasing anabolic effects on muscles thus having an association with age-related declines in the maximal voluntary neuromuscular performance capacity in aging people.     

Platelet-activating factor receptor and respiratory and metabolic responses to hypoxia and hypercapnia

Activation of the platelet-activating factor receptor (PAFR) regulates neural transmission. A PAFR blocker reduced the peak hypoxic (pHVR) but not hypercapnic ventilatory (HCVR) responses in rats [Am. J. Physiol. 275 (1998) R604]. To further examine the role of PAFR in respiratory control, genotype-verified PAFR -/- and PAFR +/+ adult male mice underwent hypoxic and hypercapnic challenges. HCVR was similar in the two groups (p-NS). However, pHVR was significantly reduced in PAFR -/- mice (38 +/- 13% baseline [S.D.]) compared to PAFR +/+ mice (78 +/- 16% baseline; P < 0.001, ANOVA), with reduced tidal volume recruitments during pHVR. In addition, hypoxic ventilatory depression was attenuated in PAFR -/- mice (P < 0.01), and was primarily due to attenuation of the time-dependent decreases in oxygen consumption during sustained hypoxia (P < 0.01). Thus, PAFR expression/function modulates components of the acute ventilatory and metabolic adaptations to hypoxia but not to hypercapnia. Imbalances in PAFR activity may lead to maladaptive regulation of the tightly controlled metabolic-ventilatory relationships during hypoxia.